Lab QA Lab Practice

7/28/2019 Lab QA Lab Practice

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Laboratory Quality Assurance Program

College of Physicians & Surgeons of Saskatchewan

LaboratoryGuidelinesGeneral

Anatomic Pathology

Chemistry

Hematology

Transfusion Medicine

2013 Edition


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Laboratory Guidelines 2013 Edition Page 1

LABORATORYGUIDELINES-2013

SUMMARY OF CHANGES

The following GUIDELINES have been revised, added, or deleted in this edition of the document.

REVISED:

Reference Textbook List for Laboratories

Urinalysis

Retention Guideline

eGFR

Differential Quality Control

Procedure for WBC Estimate

ADDED:

International Sensitivity Index (ISI) Verification

Verifying or Establishing a Normal Reference Range for Routine Coagulation Testing

DELETED:

Performance of Whole Blood Glucose Testing

D-dimer

Semen/Sperm Analysis


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Table of Contents

GeneralRetention Guideline 4

Recommended Reference Textbook List for Laboratories 7

Anatomic PathologySpecimens Exempt from all Gross &/or Microscopic Pathology Laboratories 10

Surgical Pathology Reports 12

Removal of Tissue, Blocks or Slides from the Original Hospital Site 13

Performance of Non-Gynecological Cytology 15

Follow-up Reports for Gynecological Cytology 16

Follow-up Program for Cytology 17

ChemistryEstimated Glomerular Filtration Rate eGFR 19

Cholesterol/Triglyceride/Lipid Testing 20

Urinalysis 21

Reporting Sperm in Urine 21Quality Control 22

Diagnosis and Monitoring of Thyroid Disease 23

Presence of Small Amounts of Albumin 26

Crosscheck/Validation Guideline for Those Facilities with Multiple Chemistry Instruments 27

Procedure/Method Statistical Work-up/Validation Study Guidelines 28

HematologyPrinciples for Hematology Practice 31

Hematology Films/Labelling of Slides 32

Morphology of Lymphocytes 33

Differential Performance and Referral Practice Guideline 35

Red Blood Cell Morphology Reporting Guideline 37

Differential Quality Control 39Smudge Cells 42

Flow Cytometry for the Diagnosis of Lymphoma/Leukemia 43

Malaria 44

Erythrocyte Sedimentation Rate (ESR) 45

3.2% Na Citrate Anticoagulant Recommended vs. 3.8% for Coagulation Studies 46

Vitamin B12 & Folate 47

Bleeding Time 49

Crosscheck Validation Guideline for Facilities with Multiple Hematology Instruments 50

Procedure/Method Statistical Work-up/Validation Study Guidelines 51

Protocol for Validation of Linearity on Automated Hematology Analyzers 53

Procedure for WBC Estimate 56

Establishing Conversion Factor for WBC Estimation 58

Procedure for Platelet Estimates 60

Establishing Conversion Factor for Platelet Estimation 62

Indirect Platelet Count 64

International Sensitivity Index (ISI) Verification 66

Verifying or Establishing a Normal Reference Range for Routine Coagulation Testing 67

Transfusion MedicineRetention of Transfusion Medicine Records 71

Procedure/Method Statistical Validation/Work-up Guidelines Trans. Med. 72


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GENERAL


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RETENTION GUIDELINE

Laboratories vary in size, facility and extent of services provided. Clinical laboratories must

maintain thorough, accessible records that can demonstrate an acceptable standard of care and

compliance with the accreditation requirements.

The Laboratory Quality Assurance Program of the College of Physicians and Surgeons urges

laboratories to retain records, materials, or both for a longer period of time than specified for

educational and quality improvement needs.

Laboratories must establish policies that meet or exceed the following minimum requirements

for retention of documents and specimens as established by professional and/or regulatory

organizations.

References include: CSTM, CSCC, CAP, CSA, ISO, CPSS


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Record Retention Storage TimeHematology Biochemistry Microbiology Cyto-

pathology

Surgical

Pathology

Accession record 1 year 1 year 1 year 2 years 2 years

Worksheets 1 year 1 year 1 year 6 months 6 months

Instrument print-outs 1 year 1 year 1 year n/a n/aPaper copy of patient reports 3 months 3 months 3 months indefinite indefinite

Quality control/PT documents 2 years 2 years 2 years 2 years 2 years

Maintenance records 2 years 2 years 2 years Life of the instrument, plus 2

years

Service records Life of the instrument, plus 2 years

Method /instrument evaluation 2 years after the method has been discontinued

Procedure Manual 2 years after procedure has been discontinued

Technologist ID & initials

log/computer

1 year 1 year 1 year 1 year 1 year

Telephone logs 3 months 3 months 3 months 3 months 3 months

Requisition 3 months 3 months 1 year 5 years 5 years

Laboratory Information SystemsRecords

- Validation records, including

transmission of results and

calculations

- Database changes

- Hardware and software

modifications

- LIS downtime and corrective action

2 years 2 years 2 years 2 years 2 years

Biomedical Waste Manifests must be retained for a minimum of 1 year


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Specimen Retention Hematology Chemistry Microbiology Cytopathology Surgical

Pathology

Peripheral Blood

Smear

-7days

Peripheral

Blood Smear

reviewed by a

pathologist

-1 yr

Semen smears

- 3 months

Bone Marrow

Slides/Reports

- 20 yrs (adults)

- 50 yrs(children)

Whole

blood/Plasma

- 24 hrs after

report has been

finalized

Body fluids

- 48 hrs after

report has been

finalized

Urine

- 24 hrs after

report has been

finalized

Whole blood,

serum &

plasma

- 48 hrs afterreport has been

finalized

Body fluids

- 48 hrs after

report has been

finalized

Urine - routine

- 24 hrs after

report has been

finalized

24hr Urines

- samples

discarded 48

hrs after report

has been

finalized

Swabs or

specimens

- 24 hrs after

report has beenfinalized

Positive Blood

Culture

- 5 days after

reporting

Gram Stain

- one week or

until final report

is sent

Ova & Parasiteslides

- one month

Slidesneg/unsatisfactory

- 5 years

Slides

suspicious/pos

- 20 years

Fine-needle

aspiration slides

- 20 years

Cytology

Consultation/

Requisition

- Indefinitely

Male fertility

slides

- 1 year

Cytology

paraffin blocks

- 20 years

Blocks & slides

- 20 yrs (adults)

- 50 yrs (children)

Autopsy

- 20 yrs

Gross specimen

- min. 8 weeks

after issue of

report

Wet Autopsy

Tissue

- 8 weeks after

issue of report

Bone Marrow

Slides/ Reports

- 20 yrs (adults)

- 50 yrs (children)

Photographic

Transparencies

indexed and kept

indefinitely


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RECOMMENDED REFERENCE TEXTBOOKLIST FORLABORATORIES

Microbiology1) Manual of Clinical Microbiology 10th Edition; American Society for Microbiology; Murray,

Patrick R.

2) Bailey & Scott's Diagnostic Microbiology 13th Edition, Mosby, Inc.; Forbes, Betty A., et al.3) Clinical Microbiology Procedures Handbook 3

rdEdition; Issenburg

Transfusion Medicine

1) Circular of Information, Canadian Blood Services (most recent version)2) Canadian Standards Association Blood and Blood Component Z902 (most recent version)3) Canadian Society for Transfusion Medicine (most recent version)4) Modern Blood Banking and Transfusion Practices 6th Edition; Harmening, Denise M.5) Canadian Medical Association Journal Guidelines for Red Blood Cell and Plasma

Transfusion for Adults and Children; supplement to CAN MED ASSOC J 1997;

156 (11)

6) American Association of Blood Banks Technical Manual - most recent edition7) Bloody Easy 3: Blood Transfusions, Blood Alternatives and Transfusion Reactions 3rd

Edition; Ontario Regional Blood Coordinating Network

Hematology1) Color Atlas of Hematology, Hematology and Clinical Microscopy Resource Committee,

CAP; Glassy, Eric F.

2) Clinical Hematology: principles, procedures, correlations 2nd edition; Stiene-Martin,Lotspeich-Steininger, Koepke

3) Hematology: Clinical Principles and Applications 4th Edition; Rodak, Fritsma, Doig4) Clinical Hematology Atlas 4th Edition; Carr/Rodak

Chemistry

1) Tietz Fundamentals of Clinical Chemistry 6th

Edition, W. B. Saunders Company; Burtis,Ashwood, Bruns


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2) Clinical ChemistryPrinciples, Procedures, Correlations 6th

Edition; Bishop

Urinalysis

1)

Graffs Textbook of Urinalysis and Body Fluids 2

nd

Edition; Mundt, Shanahan

Anatomic Pathology

1) Histotechnology: A Self Instructional Text 3rd Edition; Carson & Hladik2) Principles of Anatomy & Physiology 13th Edition; Tortora & Grabowski

Safety1) Transportation of Dangerous Goods Act and Regulations

Supplement Canada Gazette, Part II [www.tc.gc.ca/eng/tdg/clear-tofc-211.htm]

2) CSMLS GuidelinesLaboratory Safety, 7th Edition

Competency Evaluation

1) Canadian Society of Medical Laboratory SciencePO Box 2830 LCD 1Hamilton, ON L8N 3N8

websitewww.csmls.org

Certification - Competency Profiles
http://www.csmls.org/http://www.csmls.org/http://www.csmls.org/http://www.csmls.org/


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ANATOMICPATHOLOGY


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SPECIMENS EXEMPT FROM ALL GROSS &/ORMICROSCOPIC

PATHOLOGY LABORATORIES

Irrespective of the exemptions listed below, gross &/or microscopic examinations will be

performed whenever the attending physician requests it, or at the discretion of the pathologistwhen indicated by gross findings.

SPECIMEN TYPE DISCRETIONARY GROSS &/OR

MICROSCOPIC

Abdominal pannus Accessory Digits Amputation Bone or cartilage removed from the arthritic

joints during joint replacement surgery

Bone segments removed as part of corrective

orthopedic procedures (for example: rotatorcuff, synostosis repair, spinal fusion)

Bunions and hammer toes Calculi (renal bladder etc.), are sent for

chemical analysis and description

(By Chemistry Dept.)

Disc Materials Extraocular muscle from corrective surgical

procedures (strabismus repair)

Femoral head removed for prosthesis (if

straight forward)

Foreign bodies, such as bullets or

medicoloegal evidence that is given directlyto law enforcement personnel

Gangrenous and traumatized limbs Intrauterine contraceptive devices without

attached soft tissue

Loose bodies (joint) Medical devices such as catheters,

gastrostomy tubes, myringotomy tubes,

stents and sutures that have not contributedto patient illness, injury or death

Middle ear ossicles Nasal bone and cartilage from rhinoplasty orseptoplasty

Orthopedic hardware and other radioopaquemechanical devices provided there is an

alternative policy for documentation


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SPECIMEN TYPE DISCRETIONARY GROSS &/OR

MICROSCOPIC

Placentas which do not meet the criteria for

examination

Prosthetic breast implants Prosthetic cardiac without attached tissue

Rib segments or other tissue removed only

for the purpose of gaining surgical access

from patients who do no have a history ofmalignancy

Saphenous vein segments harvested for

coronary artery bypass

Skin or other normal tissue removed duringcosmetic or reconstructive procedures

(blepharoplasty, cleft palate repair,

abdominoplasty, rhinectomy or syndactyly

repair) that is not contiguous with a lesionand that is taken from a patient who does not

have a history of malignancy

Teeth without attached soft tissue Therapeutic radioactive sources Tonsils and adenoids if clinically not

suspicious

(under 10 yrs

old)

(over 10 yrs)

Torn menicus Varicose veins


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SURGICAL PATHOLOGY REPORTS

Timeliness of reports is critical to providing quality of care. Guidelines for surgical pathology

reporting include:

(i) Routine Surgical Pathology reports Complete within 2 working days. Where additional procedures are being performed, an extension of 24

hours is appropriate.

(ii) Autopsy Reports Written initial reports of gross pathological findings within 72 hours. Final Report30 days for routine cases, 90 days for complicated cases


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REMOVAL OF TISSUE,BLOCKS ORSLIDES FROM THE ORIGINAL HOSPITAL SITE

Anatomic Pathologists are charged with the responsibility of keeping and guarding the integrity

of the ever-growing number of tissue containing paraffin blocks and slides derived from surgical,

cytological and autopsy diagnostic services. Documentation and maintenance (tracking) of the

continuous care shall ensure quality practice. These specimens must be maintained in orderlyfiles to ensure ready access. There are inevitably increasing demands for slides, blocks or tissues

to be retrieved from the original site.

* A release form must be provided and retained on f il e at the original instituti on for permanent

release.

Summary Follow HIPA guidelines Ensure there is sufficient material for further work-up Indicate reason for request Return all material as soon as possible The lab is the custodian of tissue, blocks or slides collected. The source of material remains the property of the patient.

These are the various suggested categories to be considered:

In-Province Consultation

Request may be initiated by the primary physician, surgeon or oncologist for review by localor out-of-district pathologist.

Request may be initiated by the original signing out pathologist who is responsible formaintaining records and assuring return of the material.

Note: The return of materials to the original site must be documented and a consult report sent

to the original pathologist as well as the requesting pathologist.

Out-of-Province (recommended slides only)

Request from an originating pathologist to seek out-of-province consultation for diagnosticpurpose.

Research or national study groups request specimen be referred to another institution fortreatment.

If blocks are requested, cut a set and send the cut set. The originals should be maintained atthe processing site.

Educational Consultation

Requests for educational rounds should be restricted to slides; to ensure integrity of patientproperty.

Request should indicate for rounds and materials returned promptly to ensure ongoingpatient care.


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Research

Regulations require pathologists to obtain patient authorization and/or an InstitutionalReview Board (Ethics Committee) waiver of informed consent when using any identifiable

patient health information for research purposes.

Requests must ensure the integrity of the patient material. All materials that have critical diagnostic, prognostic or medical-legal implication may beretained at the discretion of the releasing institution. Return all materials as soon as possible.

References:Guardians of the Waxand the Patient. Editorial.; American Journal of Clinical Pathology 1995 104 p 356-7

Use of Human Tissue Blocks for research. Association of Directors of Anatomic and Surgical Pathology. Human Pathology1996.27 p 519-520


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PERFORMANCE OFNON-GYNECOLOGICAL CYTOLOGY

Non-gynecological cytology comprises of fine needle aspiration biopsies (FNAB) oforgans/tissues such as lungs and other visceral lesions, effusion cytology of pleural, peritoneal,

pericardial fluids, cytology of urine, CSF, sputum, broncho-alveolar lavage (BAL) and brush

biopsies of endoscopic procedures, (gastrointestinal tract, etc). Scrapings of open lesions andnipple discharges may also be included along with cytology of transplant organs to test for

rejection (kidneys), or for cyclosporin toxicity. BAL and transplant cytology is usually done in

specialized centers as it requires specific interpretation and often special tests. Most of the other

samples can be handled and processed in a routine surgical pathology/cytology lab equippedwith basic facilities including a biological safety cabinet (fume hood), cyto-centrifuge and

staining capability for H & E and PAP stains.

In contrast to gynecological cytology, non-gynecological cytology (NGC) does not necessarilyrequire a screening step. If adequate diagnostic material is present the focus is on diagnosis of

and interpretive correlation with the clinical setting. If cell block or cytospin samples are

available, further testing with special procedures could be performed.

Some aspects of NGC require a rapid turnaround time such as FNAB performed under CT-scan

or ultrasound guidance and intra-operative cytology requests.

It is important for institutions with CT scanner facilities to be able to provide cytology service in

house. However, it is acceptable, if the lab does not have a cytology department, the

technologists in the histology lab are trained in processing the specimens. Such training is easilyobtained and can be provided by short courses provided on site and documented.

Most NGC procedures are performed on patients who are in-patient residents in a hospital/healthcare institution or are required to come in for a day procedure/ambulatory care. Due to the time

factor involved in patients institution stay; a rapid turnaround time becomes a key factor in

availability of the service. On the other hand, the patient in an acute care setting may have an

infectious process or malignancy requiring rapid diagnosis and treatment.

The final interpretation and reporting of non-gynecological cytology shall be made by a

pathologist.


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FOLLOW-UP REPORTS FORGYNECOLOGICAL CYTOLOGY

To ensure a quality cytology service, a follow-up mechanism must be in place to provide reports

to the primary and/or consulting physician.

In an attempt to eliminate the potential for LOST TO FOLLOW reporting situations, thefollowing require follow-up letters to the primary and/or consulting physician:

1. A repeat smear was requested at the time of reporting and 3 months have lapsed since thedate of request.

2. A diagnosis is rendered requiring follow-up and none has occurred. For example: HSIL required follow-up a.s.a.p. and if this has not occurred within 3 months, a letter

is required.

LSIL (ASCUS, AGUS) within 6 months requires a letter in 9 months.

A malignant diagnosis with no apparent followup.

3. A follow-up letter has been previously issued with no reply. These letters should beautomatically generated by the computer system and then replies must be recorded and

reviewed quarterly.

4. A minimum laboratory requirement of a computer system used to report gynecologicalcytology shall have the ability to generate automatic follow-up letters that are linked todiagnostic codes.


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FOLLOW-UP PROGRAM FORCYTOLOGY

A follow-up mechanism shall be in place to ensure that actions appropriate for abnormal findings

are implemented.

a) All patients who are reported to have a significant abnormality should be followed up by

the laboratory or other agency to which this task may be delegated, to obtain final clinical

or preferably tissue confirmation of the diagnosis.

b) Statistical data should be maintained which would include the number of cases screened

annually in each category, and all correlative follow-up data available. Discrepancies, if

any, should be included with this information.


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CHEMISTRY


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ESTIMATED GLOMERULARFILTRATION RATE -eGFR

The glomerular filtration rate is the estimated volume of glomerular filtrate that moves from

renal glomerular capillaries into the Bowmans capsule per unit time . Evidence-based clinical

practice guidelines suggest that an estimate of GFR (eGFR) provides the best clinical tool togauge kidney function. The Canadian Society of Nephrology has recommended that laboratoriesreport eGFR routinely for adult patients, in order to detect chronic kidney disease.

Serum creatinine results can vary significantly and tend to be ineffective in general practice as anearly marker. 24 hr. collection for creatinine clearance is impractical and prone to error. eGFR

should be reported on outpatients over the age of 18 years. eGFR has not been validated for use

in hospitalized patients and therefore, is not recommended for reporting on inpatients.

eGFR is less reliable in, patients with near normal eGFR unstable serum creatinine acute illness extremes of body composition (eg. obesity, cachexia) unusual muscle mass (e.g. marked muscularity, muscle disease, amputation) pregnancy age under 18 years or over 70 years drugs with significant renal toxicity or clearance drugs affecting creatinine metabolism or clearance unusual dietary intake (e.g. vegetarians) other serious comorbid conditions

Summary:

Reporting of eGFR is becoming the standard of care in helping identify, stage and monitorpatients with chronic kidney disease.

eGFR > 60 Normal or slightly decreased kidney function (stages 1 or 2)

eGFR 30-59 Moderately decreased kidney function (stage 3)

eGFR 15-29 Seriously decreased kidney function (stage 4)

eGFR < 15 Kidneyfailure (stage 5)

eGFR is frequently used for DRUG DOSING using the Cockroft-Gault equation. eGFR-MDRD has not beenvalidated for this purpose.

eGFR-MDRD assumes steady state. For rapidly changing kidney function, monitor serum creatinine.(MDRD: Modification of Diet in Renal Disease)

The reported eGFR shall be multiplied by 1.21 for patients of African descent.NOTE: This information is intended for clinicians, patients and allied health professionals.

References: www.jasn.org, www.csnscn.ca, www.renal.orghttp://www.kidney.org/professionals/kdoqi/gfr_calculator.cfm
http://www.jasn.org/http://www.jasn.org/http://www.csnscn.ca/http://www.csnscn.ca/http://www.renal.org/http://www.renal.org/http://www.kidney.org/professionals/kdoqi/gfr_calculator.cfmhttp://www.kidney.org/professionals/kdoqi/gfr_calculator.cfmhttp://www.kidney.org/professionals/kdoqi/gfr_calculator.cfmhttp://www.renal.org/http://www.csnscn.ca/http://www.jasn.org/


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CHOLESTEROL /TRIGLYCERIDE /LIPID TESTING

Cholesterol results are based on optimal performance of testing. Considerations to be included:

a)

treatment/preparation of the patientb) the emergent or non-emergent nature of the testc) appropriate technical equipmentd) adequate quality controlThe accomplishment of treatment goals also demands accurate cholesterol measurements. This

requires standardization of all cholesterol measurement for accuracy to minimize the method-

specific biases. This can be achieved ONLY by standardizing the cholesterol measurements and

ensuring accuracy that is traceable to the National Reference System for Cholesterol(NRS/CHOL), National Cholesterol Education Program.

Clinical protocols are well established and should be followed by all testing sites.

Laboratories testing for lipids shall be capable of performing the entire profile, to include:

Cholesterol, Triglycerides, HDL and LDL, for diagnosis and assessment. All lipid

measurements should be performed by the same methodology.

Only instrumentation capable of maintaining intralaboratory precision that is less than or equal to

3 % (C.V.); and can demonstrate an accuracy bias of less than 3 % from the true value may beused for cholesterol analysis.


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URINALYSIS

Complete urinalysis shall include microscopic examination if the following are present:

leukocyte esterase, blood, protein, turbidity and nitrites.

The microscopic examination should be completed within four hours of collection. (Note:specimens not tested within 2 hours should be refrigerated.)

Report semi-quantitative results in SI units.

REPORTING SPERM IN URINE

Sperm are not normally present in urine and if presence is detected, it is usually considered acontaminant.

When sperm appear in a microscopic exam, they shall be reported as present. However, prior toreporting, results should be discussed with the attending physician, in order to avoid errors (ie.

mislabelling, etc.).


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QUALITY CONTROL (QC)

The purpose of QC is to detect the problems early enough to prevent their consequences.

QC emphasizes statistical control procedures, but may also include non-statistical checkprocedures, such as linearity checks, reagent and calibration checks, etc.

Two or three different materials should be selected to provide concentrations that monitor

performance at different levels of medical decision-making.

For quantitative tests, the use of two levels of control material shall be run each day of use, as a

minimum.

For qualitative tests that include built-in controls, a positive and negative control shall be

performed a minimum of once per month and upon initiation of a new lot number and shipment.

For those that do not include a built-in control, known positive and negative external controlsshall be tested each day of use.


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DIAGNOSIS AND MONITORING OF THYROID DISEASE

Thyroid function tests are among the most commonly ordered laboratory tests in the province. In

the past, investigations of thyroid disease required more than one test. Sensitive thyroid

stimulating hormone (TSH) is the initial test in the diagnosis of hypothyroidism (TSH elevatedor above normal) and hyperthyroidism (TSH is suppressed or below normal). Free T4 (FT4) ispreferred over total T4 (TT4) measurement to confirm the diagnosis of hypothyroidism or

hyperthyroidism. FT4 is 0.02 0.04% of total T4. FT4 is the metabolically active form of TT4

and is a better indicator of thyroid status than TT4 because it is unaffected by protein bindingabnormalities such as pregnancy and oral contraceptives. Free T3 (FT3) is mainly of value in

diagnosing T3 toxicosis, in determining the T3 response to therapy, and clarifying protein binding

abnormalities. It can also be of use in the early progression of subclinical hyperthyroidism to

overt thyrotoxicosis when FT4 is normal and TSH is suppressed. FT3 is often the first to beincreased. FT3 is approximately 0.20.5% of total T3.

American Association of Clinical Endocrinologists (AACE) recommends that the TSH referencerange run from 0.3 - 3.0, versus the old range of 0.5 - 5.5. Keep in mind that there is

disagreement among practitioners.

Limitations:1. These guidelines do not apply to neonates.2. TSH is not reliable in the investigation of hypothalamic or pituitary disease.3. TSH may be an unreliable indicator of thyroid status in patients with acute severe non-

thyroidal illness (e.g. CCU and ICU patients) and the test is only recommended when there

are clinical indicators of possible pre-existing thyroid disease.

4. Medications such as lithium, amiodarone, glucocorticoids, and dopamine affect TSH andmay also affect the individuals thyroid status.

Clinical Aspects of Testing:

1. Screening asymptomatic, apparently healthy patients for thyroid disease is not consideredindicated at this time.

2. Testing is indicated in the presence of symptoms or signs that are suggestive of thyroiddisease especially in high risk populations.

3. High-risk groups include women over 50, the ambulatory elderly, postpartum, individualswith a strong family history of thyroid disease and other autoimmune diseases such as Type I

diabetes.

Symptoms and Signs of Hypothyroidism: Cold intolerance, lethargy, depression, constipation,

menstrual disorders, dry skin, weight gain. There will be slow growth in children.

Symptoms and Signs of Hyperthyroidism: Palpitations, fatigue, weakness, increased appetite,heat intolerance, usually enlarged thyroid, weight loss, warm moist skin, tremor and tachycardia.

Restlessness, sleep disturbances, difficulty maintaining attention and concentration occur in

children.


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Recommended Testing Algorithm

If TSH screen is abnormal, a FT4 will be done on the same sample reducing the need to call back

the patient for subsequent testing. If the TSH is less than 0.3mU/L a FT4 and FT3 will be done

on the same sample. Free T3 is a better indicator of the degree of thyrotoxicosis in most patients.

Further Testing Recommendations:

1. Follow-up for Primary hypothyroidism or replacement therapy: TSH should be performed 6 8 weeks after start of therapy or dosage adjustment. After normal results have been

achieved, TSH should be done annually, unless the clinical condition changes or unless the

clinical condition warrants re-testing. In children under one year of age the TSH and FT4

should be measured every 3 months and every 6 months for children under six years of age.Also rapidly growing adolescents should have a TSH checked once every 6 months.

2. After Radioactive iodine treatment: A FT4 should be done at 4-6 weeks interval for the first 6months or until normal. At 6 months and then annually a TSH should be done to detecthypothyroidism. In most children and young people following thyroid ablation the FT3 is a

better indicator of control.

3. Antithyroid drugs for Hyperthyroidism: Patients should be monitored by means of FT4monthly until controlled and then at least every 3 months while on medication. If clinicalsigns and symptoms are present a FT3 may be indicated. In cases of T3 toxicosis a FT3

should be ordered. In patients with thyrotoxicosis the TSH may not recover for quite some

time after euthyroidism has been achieved and sometimes requires a period ofhypothyroidism before recovery.

4. Suppressive doses of thyroxine: Designed to support a neoplasm or goiter. TSH and FT4every 2 months until TSH has reached a level of suppression acceptable to the clinician.

5. Subclinical hypothyroidism: Borderline results are fairly common in elderly patients andindividuals with an autoimmune mechanism are more likely to progress to a hypothyroid

state. Observation and monitoring by TSH at 6-12 month intervals is recommended.

TSH

Increased Normal Decreased

FT4 low If clinical suspicion

is high for secondaryhypothyroidsm

order FT4 as initialtest

FT4 high FT4 normal

Primaryhypothyroidism

hyperthyroidism FT3 if clinicalsuspicion is high


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6. Subclinical hyperthyroidism: Patients with low or suppressed TSH but thyroid hormones inthe normal levels with minimal or no symptoms are followed by FT4 and/or FT3 at 6 12

months to gauge the progression of their condition.

References:1. HSURC guidelines. Nov 19922. Ontario Association of Medical Laboratories. Guidelines for the use of serum tests to detect thyroid dysfunction.

May 19873. Ontario Association of Medical Laboratories. Guidelines for the use of serum testing in the management of

primary hypothyroidism. June 19984. Alberta Medical Association. Laboratory Testing Guidelines for Investigation of Thyroid Dysfunction. May 19995. Protocol Steering Committee B.C. for the use of Thyroid function tests in the diagnosis and monitoring of patients

with thyroid disease. Aug 19976. The College of Physicians and Surgeons of Manitoba. Investigation of Thyroid Disease. February 19957. National Academy of Clinical Biochemistry. Laboratory Support for the Diagnosis and Monitoring of Thyroid

Disease. November 20008. Vanderpump MPJ, Ahlquist JAO, Franklyn JA et al 1996. Consensus statement of good practice and audit

measures in the management of hypothyroidism and hyperthyroidism. BMJ 313: 539-44.9. Singer PA, Cooper DS, Lewy EG et al 1995. Treatment guidelines for patients with hyperthyroidism and

hypothyroidism. JAMA 273: 808-12.10. Ladenson PW, Singer PA, Ain KB et al 2000. American Thyroid Association Guidelines for detection of thyroiddysfunction. Arch Intern Med 160: 1573-5.


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PRESENCE OF SMALL AMOUNTS OF ALBUMBIN

Presence of small amounts of albumin in urine is considered an early predictor of the

development of glomerular damage in the absence of overt nephropathy. Patients with diabetes

and hypertension are the primary risk groups.

The Canadian Diabetes Association recommends testing for small amounts of albumin once/year

after the onset of diabetes.

The presence of small amounts of albumin in urine is detectable by dipstick methodologies, and

is approved as a screen for renal damage in the known diabetes patient.

All positive results must be confirmed by quantitative analysis.


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Laboratory Quality Assurance ProgramCollege of Physicians & Surgeons of Saskatchewan

Laboratory Guidelines 2013 Edition Page

CROSSCHECK/VALIDATIONGUIDELINE FORTHOSE FACILITIES WITH MULTIPLE CHEMISTRY/HEMATOLOGY INSTRUMENTS

PERFORMING THE SAME TEST PROCEDURE

Proficiency testing registration is mandatory for each analytical test. Rotating proficiency testing result submission between analyzers is not a requirement, but is suggested where appropriate (e.g. Blood Gases Where there are multiple analyzers performing the same test procedure in larger facilities, it may be more appropriate for proficiency testing

submission to be consistently submitted from the same analyzer for tracking purposes. An internal cross-check/validation protocol is required to ensure that there is correlation between all analyzers providing the same test result

in the same facility.

If this protocol is not followed, then each analyzer must be registered in the external proficiency testing program as mandated by LQAP. This procedure is recommended every six months.

Chemistry/Hematology High Volume Analyzers (e.g. Electrolytes/CBC)

Validation/Crosscheck

Element:Requirement:

Frequency/Data Points:

Patient correlation Regularly scheduled intervals Whenever criteria for recalibration/validation is met:

- change of manufacturer for reagents or equivalent- after maintenance or service as per manufacturers

recommendations

- as required for purposes of troubleshooting/validation of reagent lot # changes or as indicated

by quality control data

Minimum of 20 patient specimens/2 times per year orequivalent

(i.e. 10 patient specimens/4 times per year or on-going data

collection as appropriate) As necessary per recalibration/validation event

Chemistry/Hematology Low Volume Analyzers (e.g. Fibrinogen)


Element: Requirement:Frequency/Data Points:



recommendations



Minimum of 5-10 patient specimens/2 times per year orequivalent

As necessary per recalibration/validation event


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PROCEDURE/METHOD STATISTICALWORK-UP/VALIDATIONSTUDY GUIDELINES:CHEMISTRY/HEMATOLOGY

Work-up guidelines and definitions (change in method/instrument):

Work-up

Element

Definition: CLIS or International Federation of Clinical

Chemistry (IFCC)

Minimum Data Requirements (where

appropriate):

Applicability:

Qualitative Quantitative

1. Imprecision within runbetween run

The variation in analytical results demonstrated when a particularspecimen of aliquot is analyzed multiple times or on multiple days.

Imprecision is expressed quantitatively by a statistic such as standard

deviation or coefficient of variations.

Within runuse preferably a patientsample or pool close to the decision levels

with a minimum of 10 data points.

Between run20 results from 20 separate

runs on 2 levels over a 10-day minimum

time period using appropriate QC material.

2. Patient

Correlation

The correlation coefficient is a means to look for a relationship, not

agreement, between pairs. Two methods may have a perfect

correlation throughout the measuring range but may not agree in

value (i.e. one may be double the value of the other).

40 data points are recommended with a

minimum of 20 having 50% of the data

points outside the reference intervals, if

possible. Correlations should involve

comparison with an acceptable reference

method or laboratory.

n=20

n=40

3. Linearity (IFCC) The range of concentration or other quantity in the specimen

over which the method is applicable without modification (CLIS)

when analytical results are plotted against expected concentrations;

the degree to which the plot curve conforms to a straight line is a

measure of the system linearity.

4 data points each in duplicate as a

minimum requirement, but 5 data points are

preferred (over reportable range). Linearity

studies are expected on an initial method

work-up and further studies as defined by

the College guidelines (i.e.

troubleshooting).

4. Reference

range

validation

It is common convention to define the reference range or interval of a

laboratory test as the central 95% interval bounded by the 2.5 and

97.5 percentiles of the selected patient population. Validation of an

established reference range requires a minimum of 20 samples.

The minimum requirement is 20 data points

for confirmation of an established reference

range and 120 for the establishment of a

new reference range.

5. Accuracy Closeness of the agreement between the result of a measurement and

the accepted reference value (true value of the analyte)

3 data points using acceptable reference

material (i.e. CEQAL or CAP) 1 data point

may be acceptable for haematology

accuracy studies if related to sample

stability.

6. Sensitivity Measure of the ability of an analytical method to detect small

quantities of the measured component. When concern is performance

at a very low concentration it is useful to determine the detection

limit as influenced by imprecision.

Sensitive studies are only required for those

methods which have clinical relevance at

values close to 0 (i.e. TSH)

When

clinically

relevant

When clinically

relevant


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7. Specimen

Stability

The conditions of handling and storage, which permits the

measurement and reporting of a clinically relevant result.

No data generally required. As per

manufacturers guidelines. If

manufacturers stability window is to be

extended a stability study is expected.

Storage and

transportation

dependent

Storage and

transportation

dependent

8. Interference The effect of any component of the sample on the accuracy of the

measurement of the desired analyte.

Document the manufacturers interference

information. The method should include a

disclaimer or a process for dealing with a

lipemic, icteric or hemolyzed sample.

Methods with

known

interferences

Methods with

known

interferences

9. Recovery A recovery procedure involves the addition of a known amount of

analyte to an aliquot of sample. Recovery is defined as the ratio of

the amount of the analyte recovered to amount added and is given as

percentage.

Recovery studies should only be necessary

for those methods or analytes where

organic extractions or equivalent are

required as part of the methodology (i.e.

Toxicology)

Method

specific/organi

extraction

Work-up requirements when an instrument is moved from site A to site B: (It is assumed that the instrument has been in recent u

with acceptable performance).

Work-up Element Minimum data requirements:

1. Imprecision studies, QC only As above

2. Patient correlation 10 data points, where feasible.


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HEMATOLOGY


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PRINCIPLES FORHEMATOLOGY PRACTICE

Hematology incorporates leading edge technology to help decipher and treat troubling diseases.

As hematology has evolved, there are some generic principles that must be simply stated for

quality patient care.

Quality management practices are essential to ensuring quality care. Some of the core principles

include:

1) Hemoglobin, the single most common complex organic molecule (Hb) shall be determinedby spectrophotometric methodology.

2) WBC and platelet counts shall be tested by automated methodology as part of a CBC onwhole blood specimens. If necessary, WBC and platelet counts may be estimated on the

peripheral smear.

3) Manual PT (INR)/APTT testing shall be discontinued. Please refer to CLSI H21-A4 forreference on collection and storage.

4) Reticulocyte Count shall be performed by automated methodology. Manual counts may usedas a QC method for automated analyzers.

5) All differential leukocyte counts shall be reported in absolute values. Reporting percentagesis optional, and would be in addition to absolute values.


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HEMATOLOGY FILMS/LABELLING OF SLIDES

Unequivocal patient identification is the first step to ensuring a quality hematology slide.

With no formal standards in place for labelling slides, the basic principles require: Unique identification of the patient (at least two identifiers) Written instructions for labelling Labels shall be clear and legible Date of collection

CLSI Document H-20-A states: Label the samples uniquely.

Positive Patient ID

_______________________________ _______________________________Last Name First Name

PHN______________________________________________________________Date of Birth

Unique ID #

Date of Slide___________________________________


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MORPHOLOGY OF LYMPHOCYTES

Benign versus Malignant

A variety of diseases and disorders may produce changes from the normal in numbers and/ormorphology and functions of one or more of the leukocytes. The most important feature ofvariant lymphocyte morphology is the recognition of its benign nature. The pertinent fact is that

these lymphocytes are normal cells that have been altered as the result of a normal response to

stimulus. When changes in WBCs are produced by non-malignant disorders (e.g. infections),the cells formerly called atypical, are now referred to as reactive lymphocytes. When changes

are suspicious of being produced by malignant disorders (leukemias, lymphomas,

gammopathies) and additional investigations may be required, the cells are often referred to as

atypical and/or abnormal.

In non-malignant disorders, the variant lymphocytes, reactive lymphocytes, atypical

lymphocytes, virocytes, stress lymphocytes, Downey cells, transformed lymphocytes,transitional lymphocytes, and glandular fever cells, among others, are normal cells reacting to a

stimulus, whether it be viral or other. The designation of reactive lymphocytes is preferred.

In Chronic Lymphocytic Leukemia, the lymphocytes are somewhat larger than normal, havenuclei with clumped or condensed chromatin, and may have prominent nucleoli. The cytoplasm

may be abundant, nongranular and moderately basophilic, or it may be relatively scant.

In Prolymphocytic Leukemia, the prolymphocyte is a relatively large mononuclear lymphoid cell

with an oval to round nucleus, coarse-appearing chromatin strands and one or two large vesicular

nucleoli with perinuclear condensations of chromatin. The cytoplasm is abundant and usually

granular and is basophilic with Romanowsky stains.

In Waldenstrms Macroglobulinemia, the abnormal B-lymphocytes involved are transitional

cells. They have the ability to differentiate into large plasmacytoid lymphocytes and plasmacells. These malignant cells circulate in the peripheral blood only in the terminal stages.

In Lymphomas, peripheral blood involvement (i.e., abnormal circulating cells) is seen late in the

disease. Lymphoma cells can exhibit a variety of appearances and the cellular morphology isvariable and depends on the underlying type of lymphoma. These cells can exhibit variable size,

shape, nuclear, and cytoplasmic characteristics. Lymphoma cells are usually round to oval, and

can be irregular. Cell size ranges from 8 to 30 m and the N-C ratio varies from 7:1 to 3:1. Indiffuse small lymphocytic lymphoma (the tissue equivalent of chronic lymphocytic leukemia),

the cells are generally small with round to oval nuclei, compact and coarse chromatin, and have a

scant amount of basophilic cytoplasm. They may be the same size as normal lymphocytes or

may be slightly larger. Occasionally, the nuclei exhibit an angulated appearance with slightlymore open chromatin. A small nuclear indentation may be present. Nucleoli are not seen.

Scattered prolymphocytes, which are larger cells with a centrally placed nucleus, a prominent

single nucleolus, and moderate basophilic cytoplasm, often are seen. In the small-cleaved celllymphomas, the cells are slightly larger than normal lymphocytes and have an angulated


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appearance. The majorityof nuclei have clefts, indentations, folds, convolutions, and may even

be lobulated. The chromatin is moderately coarse and one or more nucleoli may be prominent.

Their cytoplasm is scant to moderate and basophilic. The cells in small noncleaved lymphomas(Burkitts lymphoma) appear similar to L3 lymphoblasts. These cells are generally moderate in

size (10 to 25 m) and have a round to oval nucleus with moderately coarse chromatin, and one

or more prominent nucleoli. The cytoplasm is moderate, stains dark blue, and may containnumerous small vacuoles. Large cell lymphomas and immunoblastic lymphomas may exhibitsome of the most blast-like and abnormal morphology. These cells are large (20 to 30 m) and

have scant to moderate amounts of deeply basophilic cytoplasm. The nuclei are generally round

to oval, but may be angulated, folded, indented, or convoluted. Nucleoli are prominent and maybe single or multiple. Vacuoles can occasionally be seen in the cytoplasm. These cells can be

easily confused with blasts. T cell lymphomas can exhibit similar morphology to any of the

above types of lymphomas. The typical appearance is a moderate-size cell with a markedly

convoluted nucleus giving a cerebriform or grooved pattern. Their chromatin is moderatelycoarse and nucleoli are not apparent. The cytoplasm is generally scant and blue.

In Hairy Cell Leukemia, the abnormal lymphocytes (Hairy Cells) have scant to abundant,agranular, light grayish-blue cytoplasm. The plasma membrane appears irregular with hair-like

or ruffled projections, which are seen more easily with phase microscopy. These cells often have

a round or oval nucleus; sometimes, the nucleus appears folded or bilobed. The chromatin is

loose and lacy, and one or two nucleoli are commonly seen.

In Szary Syndrome, the abnormal lymphocyte is larger than normal with scanty cytoplasm, and

the nucleus is large with clefting. Nuclear folding can be so extensive as to suggest an image ofthe brain, and these nuclei are thus described as cerebriform. The nuclear chromatin is fine with

little condensation. There may or may not be visible nucleoli.

Note: Performance of manual differentials is required when abnormalities/unexpected results arefound in the WBC.

References:McTaggart Bill, SAIT Hematology Updating Correspondence Course, 5 th Edition, 1993. pp. 10.Stiene-Martin, 1998, pp. 355-356, 484-485, 490, 507-508. College of American Pathologists, Surveys, Hematology Glossary,2001


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DIFFERENTIAL PERFORMANCE AND REFERRAL PRACTICE GUIDELINE

Blood Film Reference Range Perform a Differential or Scan

on First Occurrence or

Significant Change

Referral

Unexpected or Unexplaine

WBC Countadults 4.0 - 11.0 x 109/LLower referral range 25.0 x 10

9/L

children (2-14 years) 5.015.0 x 109/Lchildren (90 days2

yrs)

5.020.0 x 109/L

newborn (090 days) 7.020.0 x 109/L

Absolute Neutrophils 1.57.5 x 109/L 0.5 x 109/L (all age groups)

Absolute Lymphs 1.14.4 x 109/L

adults >5.0 x 109/L >7.0 x 109/Lchildren (0-14 years) >7.0 x 109/L >10.0 x 109/L

Absolute Monocytes 0.20.8 x 109/L >1.0 x 109/L (all age groups) >1.5 x 109/L (all age groups

Hemoglobin

Lower referral rangeadult female 120160 g/L


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Blood Film Reference Range Perform a Differential or Scan

on First Occurrence or

Significant Change

Referral

Unexpected or Unexplaine

MPV 7.410.4 fL NoneLower referral range 14.0 fL

Platelet Count 150400 x 109/LLower referral range 600 x 109/L

WBC Morphology > 10% Reactive Lymphs

Pelger-Huet anomalyHypogranulated neutrophils

Hairy CellsBlasts/Immature CellsHypersegmented neutrophils

NUCLEATED RBCS >5 NRBC/100 WBC

RBC Morphology RBC inclusions: PappenheimeHowell-Jolly or Heinz Body,

basophilic stipplingPresence of schistocytes,echinocytes, bite cells, sicklecells, rouleaux,autoagglutination, significant

polychromasia, oval or round

macrocytes, target cells, teardrops, spherocytes, elliptocyteacanthocytes, stomatocytesDimorphic pictureParasitesMalaria

PLATELET

MORPHOLOGY

none

OTHERCRITERIASpecified Instrument

Flags

When indicated

Ordered by Physician Physician request Physician request

TechnologistDiscretion

Technologist initiated Technologist initiatedifsuspicious cells are present, reto a pathologist

The Color Atlas of Hematology, Hematology and Clinical Microscopy Resource Committee,

CAP; Glassy, Eric F. is recommended as a resource.

NOTE: This table is provided as a guideline only. Consult a larger centre for more specificranges.


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RED BLOOD CELL MORPHOLOGY REPORTING GUIDELINE

Red Blood Cells/100x oif Abnormality to be

Reported

Implications for Diagnosis

(200 RBC field) x 10 fields

Any present

Schistocytes/Helmet Cells Thrombotic thrombocytopenicpurpura (TTP), RBC

fragmentation syndromes such

as hemolytic uremic syndrome,DIC, microangiopathic

hemolysis, malignant

hypertension, eclampsia,

Cardiac valve hemolysis, somerenal vascular diseases

Echinocytes/Burr Cells Kidney Disease

Bite Cells Drug or chemical inducedoxidative damage, unstablehemoglobins

Sickle Cells Sickle Cell Anemia,

Hemoglobin SC/SD Disease

Basophilic Stippling Lead Poisoning, Thalassemia,

Sideroblastic & Megaloblastic

Anemia, Sickle Cell Anemia

Howell Jolly Bodies Megaloblastic Anemia, Post-

splenectomy state

Dimorphic Hemorrhage, response to

treatment, Sideroplastic anemia,post-transfusion

RBC Rouleaux (5cellsstacked)

Paraproteinemia, increasedfibrinogen, inflammatory

disorders

RBC Aggutination Autoimmune Hemolytic Anemia

(Cold Agglutinin Disease)

Parasites Identify specific forms

Nucleated RBC Sever hemolysis, part ofLeukoerythroblastic picture,

Bone marrow stress

>5 Polychromasia Response to treatment, bloodloss, Hemolysis

Macrocytes (oval) Megaloblastic state, Aplastic

Anemia, MyelodysplasticSyndrome


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Target Cells Liver Disease, Post-

splenectomy/hyposplenism,Hemoglobinopathy,

Thalassemia

Tear Drops Myelofibrosis, Pernicious

AnemiaSpherocytes Hereditary Spherocytosis,

Autoimmune Hemolytic Anemia

Pappenheimer Bodies Sideroblastic Anemia, Chronic

Hemolysis, Liver Disease

>10 Macrocytes (round) Liver Disease, Alcoholism

Elliptocytes Hereditary Elliptocytosis

Acanthocytes/Spur Cells Post-splenectomy state, Liver

Disease, Abetalipoproteinemia

Stomatocytes Liver Disease

Microcytes (hypochromiccells)

Iron Deficiency, Thalassemias,Treated Polycythemia

Avoid using the terms anisocytosis and/or poikilocytosis since they convey no specific meaning.

The numeric value is meant for in ternal use to indicate a signi fi cant abnormal ity presence. No

numeri c value is reported, just the abnormali ty.


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DIFFERENTIAL QUALITY CONTROL

The following is a list of measures undertaken by each laboratory to ensure the quality of

differential results reported on patients.

Good Laboratory Practice:

1. Develop a protocol for determining if a manual differential is required based on instrumentcapabilities. (i.e. Hematology GuidelinesDifferential Reporting Guidelines)

2. Develop a list of abnormalities, which must be reviewed by the supervisor or pathologistbefore results are reported. Refer to Blood Film Guideline.

3. Differentials must be repeated if each cell does not meet the limits set out in the table below.If the repeat is still not within the established limits, a second technologist should repeat the

differential.

4. Whenever the tech1 has concerns with a differential the rest of the CBC can be released witha notation Differential to follow. The requesting physician can be invited to review the

smear if they so choose. The smear should be evaluated as soon as possible.

5. Leukocyte abnormalities seen during a smear review require a manual differential completedregardless of the protocol for when to perform a manual differential.

6. Perform the manual differential and compare it to the automated differential using the 95%confidence limits table. Each cell should compare within the range set.

7. Differentials between technologists should also fall within the established limits (95%confidence limits).

8. If the manual differential performed by two technologists agrees within the establishedlimits, but is not in agreement with the automated differential, then the manual differential

should be reported out instead of the automated differential.

9. It is good laboratory practice to circulate unknown QA slides quarterly. The results will becompared with peers and should be within the 95% confidence limits as set by the following

table. Any problem areas will be covered between the technologist and the supervisor.

Procedure:

How to use the following 95% confidence limits table:1. Look up each cell number you want to compare to in column a.

E.g. 25% neutrophils

1 Tech refers to the medical laboratory technologist or combined laboratory and x-ray technologist.


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2. If a 100 cell differential was performed go to the column n=100 to determine theacceptable range. If a 200 cell differential was done refer to column n=200.

E.g. you counted 32 neutrophils in a 100-cell differential.

3. Determine acceptability of the differential by checking if the number you counted for acertain cell falls within the stated range.E.g. Stated range is 16-35%; you are within this range.

4. Repeat for each cell type.5. Determine acceptability of each cell line by comparing automated to manual differential. For

the neutrophils/granulocytes, segmented neutrophils, band neutrophils and other neutrophil

precursors must be added together and for lymphocytes, reactive lymphocytes and

lymphocytes must be added together.E.g. 77 segs and 15 bands = 92%

6. You must be within this range, or the differential must be repeated.

Example:

The automated or technologist differential indicated the following differential and the acceptable

range for each number was looked up in n=100 column:Neu: 70 acceptable range 60-79

Lymph: 15 8-24

Mono: 7 1-12Eos: 3 0-9

A 100 cell differential is completed by another technologist and based on the acceptable range

the results are as follows:

Neu: 52 not within limits

Lymph: 27 not within limitsMono: 12 within limits

Eos: 7 within limits

Baso: 2 within limits

This manual differential would have to be repeated.


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The 95% confidence limits for various percentages of leukocytes of a given type as determined

by differential counts on stained blood smears.

n n=100 n=200 n=500 n=1,000

0 0-4 0-2 0-1 0-1

1 0-6 0-4 0-3 0-22 0-8 0-6 0-4 1-43 0-9 1-7 1-5 2-5

4 1-10 1-8 2-7 2-6

5 1-12 2-10 3-8 3-76 2-13 3-11 4-9 4-8

7 2-14 3-12 4-10 5-9

8 3-16 4-13 5-11 6-10

9 4-17 5-14 6-12 7-1110 4-18 6-16 7-13 8-13

15 8-24 10-21 11-19 12-18

20 12-30 14-27 16-24 17-2325 16-35 19-32 21-30 22-2830 21-40 23-37 26-35 27-33

35 25-46 28-43 30-40 32-39

40 30-51 33-48 35-45 36-4445 35-56 37-53 40-50 41-49

50 39-61 42-58 45-55 46-54

55 44-65 47-63 50-60 51-59

60 49-70 52-67 55-65 56-6465 54-75 57-72 60-70 61-68

70 60-79 63-77 65-74 67-73

75 65-84 68-81 70-79 72-7880 70-88 73-86 76-84 77-83

85 76-92 79-90 81-89 82-88

90 82-96 84-94 87-93 87-9291 83-96 86-95 88-94 89-93

92 84-97 87-96 89-95 90-94

93 86-98 88-97 90-96 91-95

94 87-98 89-97 91-96 92-9695 88-99 90-98 92-97 93-97

96 90-99 92-99 93-98 94-98

97 91-100 93-98 95-99 95-9898 92-100 94-100 96-100 96-99

99 94-100 96-100 97-100 98-100

100 96-100 98-100 99-100 99-100

References:1. Abbott training seminarthe following table was provided.2. Clinical Hematology Principles, Procedures, Correlations, Cheryl A Lotspeich-Steininger et al, 1992.3. FHHR protocol


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SMUDGE CELLS

Distinguished by their naked amorphous nuclear chromatin material, smudge cells were initially

described as white blood cells with broken-down nuclei in patients with chronic lymphocytic

leukemia. Subsequently, these nuclear shadows have most often been referred to as smudgecells, but the term basket cells is used synonymously.

The mechanism is often associated primarily with traumatic disruption of cells during blood film

preparation. In the process, the cell membrane ruptures and when viewed under a microscope,what remains looks like a smudge, hence the term, smudge cells.

To ensure reliability of results, it is important to understand the effects of variables associated

with smudge cell formation, particularly the blood film preparation. Thus, the angle and thedegree of incline of the slide spreader, the type of slide spreader (sharp or smooth), the

cleanliness of the slides, and the overall quality of the blood films cannot be overemphasized.

For minimal morphologic alterations, blood films should be made within three hours and notmore than twelve hours after collection.

It is recommended to include smudge cells in the differential as an absolute count, especially

when the smudge cell numbers are noticeably increased. This identifies a more appropriatecount because smudge cells are actually lymphocyte artifacts. It also avoids the need for

repeating or verifying abnormal counts by the time - consuming albumintreated method.

Education is needed (for the ordering physicians especially) to eliminate the risk of

misinterpreting this smudge cell count as a new cell type.

Criteria for Reporting Smudge Cells

Absolute lymphocyte count should be greater than 5.0 x109/L.

Patient age should be more than 30 years*.

Smudge cells should be reported if greater than 10 per 100 leukocytes. Report smudge cells inabsolute numbers.

*Although CLL is not often diagnosed in patients under the age of 40, patients over 30 years of age should be

considered potentially at risk. CLL is rare in patients under 30 years of age.

*In children (18 & under), the smudge cells are not counted as part of the differential. However, the presence of

smudge cells may be noted on the report.

*Smudge cells are present in those candidates for which definitive diagnostic criteria are well established. Examples

include: CLL & Acute Leukemia.


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FLOW CYTOMETRY FOR THE DIAGNOSIS OF LYMPHOMA/LEUKEMIA

Immunophenotypic analysis of hematological malignancies is crucial for accurate diagnosis and

classification of these complex malignancies. Flow cytometric data shall be interpreted in the

context of additional information and correlation with other clinical findings, obtained throughgenetic studies, and through conventional morphologic and cytochemical methods. Flowcytometry by itself does not provide enough information for diagnosis.

Flow cytometric analysis has become an acceptable medical practice in the diagnosis andcharacterization of hematologic neoplasia and its role in the management of patients with these

diseases is well recognized. Despite its extraordinary power, there is great concern regarding

the inconsistent practices and wide variation in styles among laboratories involved in the flow

cytometric analysis of leukemias and lymphomas. Of particular importance are the deficienciesin standardization and validation of procedures used in the analysis, the manner by which the

information is generated and reported to pathologists or treating physicians, and the

appropriate utililzation of this technology in patient care. (US-Cdn ConsensusRecommendations on the Immunophenotypic Analysis of Hematologic Neoplasia by Flow

Cytometry)

a) A MLT with Hematology experience, and appropriate Immunology background, and

training in Flow Cytometry is required for the performance of Flow Cytometry clinical

testing of Lymphoma and Leukemia.

b) A qualified Pathologist/Hematologist who has training in both Hematopathology and

Flow Cytometry must perform interpretation of Flow Cytometry results.

c) Flow cytometry in diagnostic purposes for Lymphoma/Leukemia shall only be performed

in pathologistdirected laboratories.


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MALARIA

Malaria can be a life threatening infection and a rapid diagnosis is necessary to institute

appropriate management in a timely manner. The diagnostic information required for patient

management is best obtained by microscopy; however, a rapid test (immunologically-based) is avery useful screening test. The Rapid Diagnostic Tests (RDT) results shall be confirmed bymicroscopy.

Rapid Diagnostic Tests (RDTs):1. Should be available in all acute care sites in Saskatchewan2. A single RDT should be recommended for use across the province.3. Standardized reporting terminology should be used.4. The test should be handled as a STAT procedure.5. Results should be phoned to the requesting physician.6. All requests should automatically trigger microscopic examination.7.

Results in a patient suspected of falciparum infection should include prompt referral (ofthe patient or transport of sample) to a centre where microscopy is available on an urgent

basis.

8. Reports must indicate that the RDT is a screening test, with confirmation by microscopyto follow.

Microscopic Diagnosis:

1. The test should be available 24/7.2. Upon receipt of specimens at the testing laboratory, TAT is 4-6 hours.3. The testing should be based on the protracted (20 min.) examination of four thick and

four thin blood smears.

4. Positive slides must be confirmed by a laboratory physician and/or pathologist.5. The information in the report should include the following; a) positive or negative, b)

speciation, c) quantification of parasitemia.

6. All positive results should indicate the requirement to report Malaria cases to PublicHealth and a recommendation to consult with an Infectious Disease specialist as soon as

practically possible.

7. Results must be called to the physician by the testing laboratory8. The laboratories must participate in an external/internal quality assurance program.


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ERYTHROCYTE SEDIMENTATION RATE (ESR)

The ESR is a commonly ordered test for the assessment of inflammation and is not meant to be

used to screen an asymptomatic person for disease.

Although relatively easy to perform it is not easily monitored by conventional QC method.Therefore, the following practice is recommended.

Perform at least one ESR at two separate times. Perform within the 4 hr timeframe from collection; Results must be comparative

i.e. If the first results fall within the reference range, the second result shouldbe similar. Significant variations indicate inaccuracy and testing should be

discontinued until resolved.

Perform this practice every six months.ESR is indicated in the diagnosis and therapeutic monitoring of temperal arteritis and

polymyalgia rheumatica. The ESR may be helpful in resolving conflicting clinical evidence inpatients with rheumatoid arthritis and with the evaluation and management of patients with

specific autoimmune inflammatory or infectious disorders (e.g. pelvic inflammatory disease,

bacterial endocarditis, septic arthritis and osteomyelitis).


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3.2% Na CITRATE ANTICOAGULANT RECOMMENDED VERSUS 3.8%

FORCOAGULATION STUDIES

The International Standards Committee on Thrombosis and Hemostasis, and the CLSI have

recommended guidelines to standardize whole blood collection to 3.2% sodium citrateanticoagulant.

Several publications have indicated that the clotting times for PT and APTT are significantly

shortened with the 3.2% NaCitrate. For this reason, exchange between the two concentrations is

not acceptable. When selecting an anticoagulant for collection, it is important to note that 3.2%

NaCitrate is used for ISI assignments for thromboplastin. For these reasons, it is important forlaboratories to standardize the choice of anticoagulant for sample collection.

The anticoagulant used for coagulation assays should be 3.2% trisodium citrate. The proportion

of blood to anticoagulant volume is 9:1. Inadequate filling of the collection device will decrease

this ratio, and may lead to inaccurate results. The manufacturer recommendations should always

be followed.

If the hematocrit is very high and the usual relative volumes of blood and citrate solution are

mixed, a prolonged prothrombin time results from overcitration of the reduced proportion of

plasma in the blood sample. This means the final citrate concentration in the blood should beadjusted in patients who have hematocrit (PCV) values above 0.55 (55%). Adjust the volume of

NaCitrate in the draw tube by applying the calculation outlined in the CLSI H21-A3. Note: To

maintain the vacuum in the NaCitrate collection tube, use a tuberculin syringe to draw out theanticoagulant.

The anticoagulant volume may be calculated from the expression:

X = (100PCV) x draw volume of tube (mLs)(595PCV)

X = the volume of anticoagulant required to prepare volume of anticoagulated blood

PCV = packed cell volume (Hct) in %

For example: To determine the volume required for 5 mLs anticoagulated blood, calculate x 5

mLs

References:H21-A3 CLSI, BD Vacutainer Systems, Azko Nobel

Assessment of the influence of citrate concentration on the international Normalized Ratio (INR) determined with twelvereagent-instrument combinations. Chantarangkul, Tripodi, Clerici, Negri, Mannucci

Effect of concentration of trisodium Citrate Anticoagulant on Calculation of the international Normalized Ratio and theInternational Sensitivity Index of Thromboplastin, Duncan, Casey, Duncan, and Lloyd

The Prothrombin Time Test: Effect of Varying Citrate Concentration, Ingram, Hills.

Comparison of 3.2% vs. 3.8% Sodium Citrate Anitcoagulant Collection Tubes for Coumadin, Heparin, Abnormal and NormalSpecimens. Phillips, Sunnybrook ON


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VITAMIN B12

&FOLATE

Background:

The normal proliferation of cells depends on adequate folate and vitamin B12. Folate is

necessary for efficient thymidylate synthesis and production of DNA. B12 is needed tosuccessfully incorporate circulating folic acid into developing RBCs retaining folate in the RBC.

MEASURING BOTH B12AND FOLATE LEVELS IS NOT NECESSARY IN ALL PATIENTS.

Serum B12:The clinical indications for ordering serum B12 include:

1) Evaluation of patients with MACROCYTIC (high MCV in the CBC) anemia and the clinicalinformation suggesting possible B12 deficiency;

2) Evaluation of patients with psychiatric and neurologic impairment (symptoms of subacutecombined degeneration of spinal cord).

Red Cell Folate:

Red cell folate is ordered as it is an indication of the folate status over a longer period of time

(several months) as opposed to serum folate that reflects levels over the last few days.

Population studies have shown that dietary supplements have increased average folate levels andtherefore folate deficiency is much rarer. The clinical indications for ordering red cell folate

include the evaluation of MACROCYTIC (high MCV in the CBC) anemia and the clinical

information suggesting possible folate deficiency.

Practice Tips:

1. Persons on B12/folate supplements or other multivitamins do not require testing.2. Lab tests are used to confirm a specific diagnosis and not as a fishing expedition.3. Marginal B12 deficiency in any elderly patient with dementia, peripheral neurological

symptoms or impaired immunity should be taken seriously.

4. B12 should not be done on patients on oral contraceptives.

Prepared by Dr. A. Saxena,Department of Laboratory Medicine,Saskatoon Health Region


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Current Indications for B12 and Folate Investigation

Measuring both B12 and Folate levels is not necessary in all patients.

Notes:*B12 tests should not be done if the person is on B12

*False low B12 in pregnant women and women on oral contraceptives

Serum folates are not a useful screening toolFolic Acid Deficiency is rare due to folate fortification in food

RBCFOLATE IS MORE INDICATIVE OF TISSUE FOLATE LEVELS

Folate tests will not be done if the person is on folate

Normal

CBC

MCV

No further testingYES

B12 and RBC

folate

AbnormalNormal

Neurological or Clinicalsymptoms suggestive of B12

&/or Folatedeficiency

Requires further

patient evaluation

No further

testing


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BLEEDING TIME

Bleeding Time (BT) is defined as the time between the making of a small incision and the

moment the bleeding stops. It indicates how well platelets interact with blood vessel walls to

form blood clots (a test of platelet and vascular function).

Purpose:

Bleeding time is used most often to detect qualitative defects of platelets, e.g. Von Willebrands

disease (vWD). However, it is prolonged in many situations including vascular disorders, e.g.Ehler-Danlos syndrome, pseudoxanthoma elasticum.

Issue:

The use of BT is declining and at many institutions this test has been eliminated. This is becauseof three main reasons:

It is neither a sensitive nor a specific test.

A specific diagnosis of vWD can be made based on history and other tests.

More reliable information about platelet function can be obtained by other tests.Notes:

Use of bleeding time is not recommended (not by itself). Current practice varies and if BT is

required, the recommended method is the template device. BT is generally not recommended for

children under the age of 5.

Reference Interval:

Results are reported to the nearest half-minute. The reference range varies and is usually 1.5-9.5minutes. The test should be discontinued if the patient hasnt stopped bleeding by 15-20 minutes.

References:Andrew M, Paes B, Bowker J, et al, "Evaluation of an Automated Bleeding Time Device in the Newborn,"Am J Hematol, 1990,

35(4):275-7.Basili S, Ferro D, Leo R, et al, "Bleeding Time Does Not Predict Gastrointestinal Bleeding in Patients With Cirrhosis,"J Hepatol,1996, 24(5):574-80.Brown BA, Hematology: Principles and Procedures, 6th ed, Philadelphia, PA: Lea and Febiger, 1993, 267-70.

de Rossi SS and Glick MG, "Bleeding Time: An Unreliable Predictor of Clinical Hemostasis,"J Oral Maxillofac Surg, 1996,54(9):1119-20.

George JN, Shattil SJ: The Clinical Importance of Acquired Abnormalities of Platelet Function. NEJM 324:27-29, 1991.Gewirtz AS, Miller ML, and Keys TF, "The Clinical Usefulness of the Preoperative Bleeding Time,"Arch Pathol Lab Med, 1996,120(4):353-6.

Henry, J. B. Clinical Diagnosis and Management by Laboratory Methods. Philadelphia: W. B. Saunders Co., 1996.Munro J, Booth A, and Nicholl J, "Routine Preoperative Testing: A Systematic Review of the Evidence,"Health Technol Assess,1997, 1(12):1-62.

Lewis SM, Ban BJ, Bates I. Dacie and Lewis Practical Haematology. 2006. Churchill Livingstone Elsevier, Philadelphia (PA).Lind SE: Prolonged Bleeding Time. Am J Med 77:305-312, 1984.Peterson P, Hayes TE, Arkin CF, et al, "The Preoperative Bleeding Time Test Lacks Clinical Benefit,"Arch Surg, 1998,133(2):134-9.

Rodgers RP and Levin J, "A Critical Reappraisal of the Bleeding Time, "Semin Thromb Hemost, 1990, 16(1):1-20.Triplett DA. Laboratory Evaluation of Coagulation, 1982. American Society of Clinical Pathologists Press, Chicago (IL).


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CROSSCHECK/VALIDATIONGUIDELINE FOR THOSE FACILITIES WITH MULTIPLECHEMISTRY/HEMATOLOGY INSTRUMENTS

PERFORMING THE SAME TEST PROCEDURE

Proficiency testing registration is mandatory for each analytical test. Rotating proficiency testing result submission between analyzers is not a requirement, but is suggested where appropriate (e.g. Blood

Gases).

Where there are multiple analyzers performing the same test procedure in larger facilities, it may be more appropriate for proficiencytesting submission to be consistently submitted from the same analyzer for tracking purposes.

An internal cross-check/validation protocol is required to ensure that there is correlation between all analyzers providing the same testresult in the same facility.

If this protocol is not followed, then each analyzer must be registered in the external proficiency testing program as mandated by LQAP. This procedure is recommended every six months.

Chemistry/Hematology High Volume Analyzers (e.g. Electrolytes/CBC)






recommendations



Minimum of 20 patient specimens/2 times per year orequivalent

(i.e. 10 patient specimens/4 times per year or on-going data

collection as appropriate) As necessary per recalibration/validation event

Chemistry/Hematology Low Volume Analyzers (e.g. Fibrinogen)






recommendations



Minimum of 5-10 patient specimens/2 times per year orequivalent

As necessary per recalibration/validation event


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PROCEDURE/METHOD STATISTICALWORK-UP/VALIDATIONSTUDY GUIDELINES:CHEMISTRY/HEMATOLOGY

Work-up guidelines and definitions (change in method/instrument):

Work-up

Element

Definition: CLIS or International Federation of Clinical

Chemistry (IFCC)

Minimum Data Requirements (where

appropriate):

Applicability:

Qualitative Quantitative

1. Imprecision

within runbetween run

The variation in analytical results demonstrated when a particular

specimen of aliquot is analyzed multiple times or on multiple days.

Imprecision is expressed quantitatively by a statistic such as standard

deviation or coefficient of variations.

Within runuse preferably a patient

sample or pool close to the decision levels

with a minimum of 10 data points.

Between run20 results from 20 separate

runs on 2 levels over a 10-day minimum

time period using appropriate QC material.

2. Patient

Correlation

The correlation coefficient is a means to look for a relationship, not

agreement, between pairs. Two methods may have a perfect

correlation throughout the measuring range but may not agree in

value (i.e. one may be double the value of the other).

40 data points are recommended with a

minimum of 20 having 50% of the data

points outside the reference intervals, if

possible. Correlations should involve

comparison with an acceptable reference

method or laboratory.

n=20

n=40

3. Linearity (IFCC) The range of concentration or other quantity in the specimen

over which the method is applicable without modification (CLIS)

when analytical results are plotted against expected concentrations;

the degree to which the plot curve conforms to a straight line is a

measure of the system linearity.

4 data points each in duplicate as a

minimum requirement, but 5 data points are

preferred (over reportable range). Linearit

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