General Cell Maturation CharacteristicsIMMATURE CELLS MATURE CELLSCell is large Cell becomes smaller

Nucleoli are present Absent Nucleoli

Chromatin - fine and delicate

Coarse and clumped

Nucleus - round Round, lobulated or segmented

Cytoplasm – dark blue (rich in RNA)

Light blue (less RNA) cytoplasm

High N:C (nucleus:cytoplasm) ratio

Low N:C ratio

Erythrocyte MaturationErythropoiesis – red cell production

and maturationCD34 – hematopoietic stem cell

markerErythropoiesis is regulated by

erythropoietin, which is produced by the kidneys

Strongest stimulus for erythropoietin production: tissue hypoxia

Erythrocyte Maturation

Erythrocyte MaturationMaturation Stages:Six morphological stages of erythrocyte

maturation may be identified with Wright stainNormal maturation is dependent on intake of

proper nutrients and vitamins such as folate, vitamin B12 and iron

Nomenclature:There are three nomenclatures used to describe

the six stagesRubri (proposed by the ASCP)Erythroblast (proposed by Paul Ehrlich)Normoblast (normal precursor)

Erythrocyte Maturation

Erythrocyte MaturationGeneral Guidelines:The ASCP published the following general

guidelines for identification of erythroid precursors:Progressive decrease in size and the degree of

cytoplasmic basophilia (blue color) as the cell matures

Nuclei are round or oval in the blast stage become round thereafter

Gradual increase in coarseness and condensation of the chromatin, ranging from fine in the early stages to pyknotic in the stage just before nuclear extrusion

Rubriblast (Pronormoblast)

This is the earliest erythrocyte, cell size up to 20 m,

The nuclear:cytoplasmic ratio is 8:11-3 nucleoli, nucleus has dark areas of DNAChromatin is fine and uniform and stains intenselyDeep blue cytoplasm with no granules

Prorubricyte (Basophilic normoblast)

Size up to 16 m with an N:C ratio of 4:1Centrally located nucleus with 0-1 nucleoliChromatin coarseningCytoplasm is less blue but intensely basophilic

RNA

Rubricyte (Polychromatophilic normoblast)

Size up to 12 m with an N:C ratio of 4:1Eccentric nucleus with no nucleoliChromatin shows significant clumpingHemoglobin synthesis starts in this stageThe RNA and hemoglobin give the cytoplasm a

reddish-blue color called polychromasia or polychromatophilia

Metarubricyte (Orthochromic normoblast)

This is the last nucleated erythrocyte stageSize up to 10 m with an N:C ratio of 0.5:1Eccentrtic nucleus with small, fully condensed

(pyknotic) nucleus; no nucleoliPale blue to salmon cytoplasmHemoglobin synthesis decreases

Reticulocyte (diffusely basophilic erythrocyte)

Size up to 10 mContains no nucleus but has mitochondrial and ribosomes Last stage to synthesize hemoglobinLast stage in bone marrow before release to the bloodSupravital stain is used for identificationOne of the best indicators of bone marrow functionHemoglobin continues to be produced for approx 24 hours

after exiting the bone marrow

Mature Erythrocyte

The mature erythrocyte is approximately 6-8 m in size

It is a biconcave disc and hence referred to as a discocyte

In a Wright stain, a central pale area is revealed which fades gradually into the reddish pink cytoplasm

Granulocyte Maturation

MYELOBLAST

Earliest recognizable granulocyte precursor14-20 µm; NC ratio 7:1-4:1Round/oval nucleus with fine reddish-purple

staining chromatin2-5 nucleoli; dark blue cytoplasmNo cytoplasmic granules1% of the nucleated cells in the bone marrow

PROMYELOCYTE

15-21 µm; NC ratio 3:1Round/oval nucleus with slightly coarsening

chromatin1-3 nucleoli; dark blue cytoplasmCytoplasm has large, nonspecific/primary

granules containing myeloperoxidase2-5% of the nucleated cells in the bone marrow

MYELOCYTE

First stage where the different granulocyte types can be differentiated

12-18 µm; NC ratio 2:1; round nucleus with coarse chromatin

Light blue to light pink cytoplasm; prominent golgi apparatusSpecific/secondary granules that contain hydrolytic enzymes

(e.g. alkaline phosphatase and lysozyme) are presentNonspecifc granules are present and may still stainLast stage capable of cell division

METAMYELOCYTE

10-18 µm; NC ratio 1.5:1Nucleus is indented in a kidney bean shape and has

coarse, clumped chromatinNuclear indent less than half the width of a

hypothetical round nucleusCytoplasm is pink filled with specific/secondary granulesPrimary granules are present but do not stain

BAND (STAB)

10-15 µm; NC ratio 1:2 Nucleus is C or S-shaped with coarse, clumped chromatin Nuclear indent is greater than half the width of a hypothetical

round nucleus Cytoplasm is pink and filled with specific/secondary granules Primary granules are present but do not stain Stored in the bone marrow and released when there is increased

demand Earliest stage that can be seen in the periperal blood of a

healthy person

MATURE GRANULOCYTES

NEUTROPHIL

EOSINOPHIL

BASOPHIL

10-15 µm; NC ratio 1:3

12-16 µm 10-15 µm

Nucleus has coarsed, clumped chromatin with 3-5 lobes connected by thin filaments

Nucleus is usually bilobed

Nucleus is unsegmented or bilobed

Cytoplasm contains fine lilac secondary granules

Cytoplasm contains large, bright red-orange secondary granules; contains enzymes and proteins

Cytoplasm contains dark violet to purple blue secondary granules; contains heparin and histamine