The Biotechnology Education Company · The Biotechnology Education Company ® EDVOTEK, Inc. •...

Updated

Revised

and

The Biotechnology Education Company ®

EDVOTEK, Inc. • 1-800-EDVOTEK • www.edvotek.com

EVT 001197AM

EDVO-Kit #

104& 104-Q

Size Determination ofDNA Restriction Fragments

Storage: See Page 3 for specific storage instructions

ExpERImENT OBjECTIVE:

The objective of this experiment module is todevelop an understanding of principles involved inestimating the size of unknown DNA fragments by

agarose gel electrophoresis.

This document includes instructions for both EDVOTEK Experiment # 104 and # 104-Q. Please follow instructions for the appropriate experiment. Experiment #104 is designed for DNA staining with InstaStain® Methylene Blue. Experiment # 104-Q is designed for DNA staining with InstaStain® Ethidium Bromide.

Size Determination of DNA Restriction Fragments

EVT 001197AM

�

104104EDVO-Kit #

EDVOTEK - The Biotechnology Education Company® • 1-800-EDVOTEK • www.edvotek.com

All components are intended for educational research only. They are not to be used for diagnostic or drug purposes, nor adminis-tered to or consumed by humans or animals.

THIS EXPERIMENT DOES NOT CONTAIN HUMAN DNA. None of the experiment components are derived from human sources.

EDVOTEK, The Biotechnology Education Company, and InstaStain are registered trademarks of EDVOTEK, Inc.. Ready-to-Load and UltraSpec-Agarose are trademarks of EDVOTEK, Inc.

Page

Experiment Components 3Experiment Requirements 3Background Information 5

Experiment procedures Experiment Overview and General Instructions 7

Agarose Gel Electrophoresis Agarose Gel Requirements for this Experiment 10 Preparing the Gel Bed 10 Casting Agarose Gels 10 Preparing the Gel for Electrophoresis 11 Loading the Samples 12

Running the Gel 13 Staining and visualization of DNA 13 Study Questions 14 Instructor's Guidelines 15 Notes to the Instructor 16 Pre-Lab Preparations 19 Experiment Results and Analysis 21 Study Questions and Answers 22

Appendices 0.8% Agarose Gel Preparation A DNA Staining with InstaStain® Methylene Blue 24 B DNA Staining with InstaStain® Ethidium Bromide 25

C Quantity Preparations for Agarose Gel Electrophoresis 26 Staining and Visualization of DNA D Method 1: InstaStain® MetBlue One-step Staining and destaining 27 E Method 2: InstaStain® MetBlue Cards 28 F Method 3: Liquid Methylene Blue Plus™ 30 G Method 4: InstaStain® Ethidium Bromide Cards 31

Material Safety Data Sheets 32

This document includes instructions for both EDVOTEK Experiment # 104 and # 104-Q. Please follow in-structions for the appropriate experiment. Experiment # 104 is designed for DNA staining with InstaStain® Methylene Blue. Experiment # 104-Q is designed for DNA staining with InstaStain® Ethidium Bromide.

Table of Contents


EVT 001197AM

�

EDVOTEK - The Biotechnology Education Company® 1-800-EDVOTEK • www.edvotek.com

FAx: (�01) �40-058� • email: [email protected]

104EDVO-Kit #

Experiment Components

Requirements

READy-TO-LOAD™ DNA SAmpLES FOR ELECTROphORESIS

A Standard DNA Fragments B Unknown DNA 1 C Unknown DNA 2

REAGENTS & SuppLIES

• UltraSpec-Agarose™ powder • Concentrated electrophoresis buffer • Practice Gel Loading Solution • 1 ml pipet • 100 ml graduated cylinder (packaging for samples) • Microtipped Transfer Pipets • DNA Stain for Standard Series 100 experiments • InstaStain® Methylene Blue • Methylene Blue Plus™ • DNA Stain for Series 100-Q experiments • InstaStain® Ethidium Bromide

• Horizontal gel electrophoresis apparatus • D.C. power supply • Automatic micropipets with tips • Balance • Microwave, hot plate or burner • Pipet pump • 250 ml flasks or beakers • Hot gloves • Safety goggles and disposable laboratory gloves • Distilled or deionized water • For gel staining with InstaStain® Methylene Blue • Small plastic trays or large weigh boats for destaining • White light DNA visualization system • For gel staining with InstaStain® Ethidium Bromide • UV Transilluminator • Photodocumentation system (optional)

Store entire experiment at

room temperature.

DNA samples do not require heating prior to gel loading.

DNA samples are stable at room temperature. However, if the experiment will not be conducted within one month of receipt, it is recommended that the DNA samples be stored in the refrigerator.

Over time, some evaporation of samples may occur. Before distributing reagents to stu-dents, check sample volumes as described in the Instructor’s Pre-Lab Preparation section.

Components & Requirements


EVT 001197AM

4

104104EDVO-Kit #


Mon - Fri 9 am - 6 pm ET

(1-800-338-6835)

EDVO-TECH SERVICE

1-800-EDVOTEK

Mon - Fri9:00 am to 6:00 pm ET

FAX: (301) 340-0582Web: www.edvotek.comemail: [email protected]

Please have the following information ready:

• Experiment number and title• Kit lot number on box or tube• Literature version number (in lower right corner)• Approximate purchase date

Technical ServiceDepartment

This document includes instructions for both EDVOTEK Experiment # 104 and # 104-Q. Please follow instructions for the appropriate experiment.

Experiment #104

Designed for DNA staining and visualization with InstaStain® Methylene Blue. This experiment requires a 0.8% gel with the following volume:

30 ml (7 x 7 cm) or 60 ml (7 x 14 cm)

Refer to Table A.1 or A.2 in Appendix A for agarose gel preparation specifi-cations.

Experiment # 104-Q

Designed for DNA staining and visualization with InstaStain® Ethidium Bro-mide. This experiment requires a 0.8% gel with the following volume:

25 ml (7 x 7 cm) or 50 ml (7 x 14 cm)

Refer to Table A.3 or A.4 in Appendix B for agarose gel preparation specifi-cations.

Online Orderingnow available

Visit our web site for information about EDVOTEK’s complete line of “hands-on” experiments forbiotechnology and biology education.

5Size Determination of DNA Restriction Fragments

104EDVO-Kit #

The Biotechnology Education Company® • 1-800-EDVOTEK • www.edvotek.com

Duplication of this document, in conjunction with use of accompanying reagents, is permitted for classroom/labora-tory use only. This document, or any part, may not be reproduced or distributed for any other purpose without the written consent of EDVOTEK, Inc. Copyright © 1989,1992,1994,1997,1998, 2000, 2004, 2007, EDVOTEK, Inc., all rights reserved. EVT 001197AM

Backg

rou

nd

Info

rmatio

n


Size determination of DNA fragments is essential to DNA mapping and analyzing restriction enzyme cleavage patterns. Restriction enzymes are endonucleases that cleave both strands of DNA at very specific sequences within DNA. The location of their cleavage sites are important for DNA fin-gerprinting, determination of genetic diseases and in formulating strategies for DNA analysis.

Agarose gel electrophoresis is a convenient analytical method for deter-mining the size of DNA molecules in the range of 500 to 30,000 base pairs. Samples of DNA are loaded into wells made in an agarose gel, which is placed in an electrophoresis chamber containing a buffer solution and electrodes. Direct current (D.C.) is applied from a power source. Since DNA is negatively charged at neutral pH, it will migrate through the gel towards the positive electrode. The agarose gel consists of microscopic pores that act as a molecular sieve that separates DNA molecules according to their size and shape. The migration rate of DNA molecules of the same shape is inversely proportional to their size. This results in the smaller DNA molecule to migrate faster through the gel. The charge to mass ratio is the same for different sized DNA molecules.

Nucleotides in DNA are linked together by negatively charged phosphodies-ter bonds. For every base pair (average molecular weight of approximately 660) there are two charged phosphate linkages. Therefore, the nega-tive charge in DNA is accompanied by approximately the same mass. The absolute amount of charge in DNA is not a critical factor in the separation process. Separation occurs because smaller molecules pass through the gel pores more easily than larger ones (i.e., the gel is sensitive to the physical size of the molecule). DNA fragment migration rate is inversely proportional to the log10 of its size in base pairs.

In this experiment, DNA fragments of unknown size and Stan-dard DNA fragments are submitted to electrophoresis. The unknown DNA fragments will migrate through the gel accord-ing to their respective sizes and relative to the Standard DNA fragments. After electrophoresis, the gel is stained and the DNA bands are visualized. The migration distances of the known and unknown fragments are measured and plotted on semi-log graph paper according to their size on the y-axis versus the mi-gration distance on the x-axis. The size of the fragments on the y-axis are expressed as the log of the number of base pairs. This allows the data to be plotted as a straight line. The DNA frag-ments of known size (Standard DNA fragments) are used to make a standard curve. The migration distance of the unknown DNA fragments are extrapolated and estimated from the standard curve.

Quick Reference:

Standard DNA fragments, which were generated by restriction enzymes are provided in this experiment. A standard curve will be made on semi-log graph paper. The following are the Standard DNA fragment sizes - length is expressed in base pairs.

23130 9416 6557 4361 3000 2322 2027 725 570


EVT 001197AM

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104104EDVO-Kit #


�Size Determination of DNA Restriction Fragments

104EDVO-Kit #



The Exp

erimen

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ExpERImENT OBjECTIVE:

The objective of this experiment module is to develop an understanding of principles involved in estimating the size of unknown DNA fragments by agarose gel electrophoresis.

LABORATORy SAFETy

1. Gloves and goggles should be worn routinely as good laboratory prac-tice.

2. Exercise extreme caution when working with equipment that is used in conjunction with the heating and/or melting of reagents.

3. DO NOT MOUTH PIPET REAGENTS - USE PIPET PUMPS.

4. Exercise caution when using any electrical equipment in the laboratory.

5. Always wash hands thoroughly with soap and water after handling reagents or biologi-cal materials in the laboratory.

LABORATORy NOTEBOOK RECORDINGS:

Address and record the following in your laboratory notebook or on a sepa-rate worksheet.

Before starting the Experiment:

• Write a hypothesis that reflects the experiment. • Predict experimental outcomes.

During the Experiment:

• Record (draw) your observations, or photograph the results.

Following the Experiment:

• Formulate an explanation from the results. • Determine what could be changed in the experiment if the experi-

ment were repeated. • Write a hypothesis that would reflect this change.

Experiment Overview and General Instructions


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DNA Samples

After electrophoresis, transfer gel for staining

Analysis on UV Transilluminator

No destaining

Analysis on White light source After destaining

100 Series (Standard)

InstaStain® Methylene Blue

100-Q SeriesInstaStain®

Ethidium Bromide

© 2

006

EDV

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K, I

nc.

Attach safety cover, connect leads to power source and

conduct electrophoresis

Load each samplein consecutive wells.

Remove end blocks, comb and submerge gel under buffer

in electrophoresis chamber

Prepare agarosegel in casting tray

Gel pattern will vary depending on experiment


ExpERImENT OVERVIEw: FLOw ChART

�Size Determination of DNA Restriction Fragments

104EDVO-Kit #



The Exp

erimen

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Individual 1.5 ml or 0.5 ml microtest tubes

• Your instructor may have aliquoted samples into a set of tubes for each lab group. Alternatively, you may be required to withdraw the appropri-ate amount of sample from the experiment stock tubes.

• Check the sample volume. Sometimes a small amount of sample will cling to the walls of the tubes. Make sure the entire volume of sample is at the bottom of the tubes before starting to load the gel.

• Briefly centrifuge the sample tubes, or tap each tube on the table-top to get all the sample to the bottom of the tube.


ABOuT ThE ELECTROphORESIS SAmpLES

Samples in EDVOTEK Series 100, 100-Q and Sci-On® Series electrophoresis experiments are packaged in individual 1.5 ml or 0.5 ml microtest tubes.


10



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AGAROSE GEL REQuIREmENTS FOR ThIS ExpERImENT

• Recommended gel size: 7 x 7 cm or 7 x 14 cm

• Number of sample wells required: 3

• Placement of well-former template: first set of notches

• Agarose gel concentration: 0.8%

pREpARING ThE GEL BED

1. Close off the open ends of a clean and dry gel bed (casting tray) by us-ing rubber dams or tape.

A. Using Rubber dams:

• Place a rubber dam on each end of the bed. Make sure the dam fits firmly in contact with the sides and bottom of the bed.

B. Taping with labeling or masking tape:

• With 3/4 inch wide tape, extend the tape over the sides and bottom edge of the bed.

• Fold the extended edges of the tape back onto the sides and bottom. Press contact points firmly to form a good seal.

2. Place a well-former template (comb) in the first set of notches at the end of the bed. Make sure the comb sits firmly and evenly across the bed.

Agarose Gel Electrophoresis

• If you will be staining the gel after electro-phoresis with InstaStain® Methylene Blue, you should be using Table A.1 or A.2 found in Appendix A.

• If you will be staining the gel after electro-phoresis with InstaStain® Ethidium Bromide, you should be using Table A.3 or A.4 found in Appendix B.

CASTING AGAROSE GELS

3. Use a 250 ml flask or beaker to prepare the gel solution.

4. Use the appropriate Reference Table for aga-rose gel preparation provided by your instruc-tor. Add the specified amount of agarose powder and buffer and swirl the mixture to disperse clumps of agarose powder.

5. With a marking pen, indicate the level of the solution volume on the outside of the flask.


104EDVO-Kit #



The Exp

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6. Heat the mixture to dissolve the agarose powder.

A. Microwave method: • Cover the flask with plastic wrap to minimize evaporation. • Heat the mixture on High for 1 minute. • Swirl the mixture and heat on High in bursts of 25 seconds until

all the agarose is completely dissolved.

B. Hot plate method: • Cover the flask with aluminum foil to minimize evaporation. • Heat the mixture to boiling over a burner with occasional swirl-

ing. Boil until all the agarose is completely dissolved.

Check the solution carefully and continue heating until the final solu-tion appears clear (like water). If "crystal" particles are visible, the agarose is not completely dissolved.

7. Cool the agarose solution to 60°C with careful swirling to promote even dissipation of heat. If detectable evaporation has occurred, add distilled water to bring the solution up to the original volume marked in step 5.

After the gel is cooled to �0°C:

• If you are using rubber dams, go to step 9. • If you are using tape, continue with step 8.

8. Seal the interface of the gel bed and tape to prevent the agarose solution from leaking.

• Use a transfer pipet to deposit a small amount of cooled agarose to both inside ends of the bed.

• Wait approximately 1 minute for the agarose to solidify.

9. Place the bed on a level surface and pour the cooled (60°C) agarose solu-tion into the bed.

10. Allow the gel to completely solidify. It will become firm and cool to the touch after approximately 20 minutes.

pREpARING ThE GEL FOR ELECTROphORESIS

11. After the gel is completely solidified, carefully and slowly remove the rubber dams or tape from the gel bed.

Be especially careful not to damage or tear the gel wells when removing the rubber dams. A thin plastic knife, spatula or pipet tip can be inserted between the gel and the dams to break the surface tension.

At high altitudes, it is recommended to use a microwave oven to reach boiling temperatures.


DO NOT POUR BOILING HOT AGAROSE INTO THE GEL BED.

Hot agarose solution may irreversibly warp the bed.

60˚C

Important Note


1�



EDVO-Kit #Th

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12. Remove the comb by slowly pulling straight up. Do this carefully and evenly to prevent tearing the sample wells.

13. Place the gel (on its bed) into the electrophoresis chamber, properly oriented, centered and level on the platform.

14. Fill the electrophoresis apparatus chamber with the appropriate amount of diluted (1x) electrophoresis buffer (refer to Table B (found in Appendix A or B) on the instruction sheet provided by your instructor).

15. Make sure that the gel is completely submerged under buffer before proceeding to loading the samples and conducting electrophoresis.


LOADING ThE SAmpLES

Samples should be loaded into the wells of the gel in consecutive order.

• For gels to be stained with InstaStain® Methylene blue, the amount of sample that should be loaded is 35-38 µl.

• For gels to be stained with InstaStain® Ethidium Bromide, the amount of sample that should be loaded is 18-20 µl.

Reminder:

During electrophoresis, the DNA samples migrate through the agarose gel towards the positive electrode. Before loading the samples, make sure the gel is properly oriented in the apparatus chamber.

+Black Red

Sample wells

–

Lane Tube

1 A Standard DNA Fragments 2 B Unknown 1 3 C Unknown 2

1�Size Determination of DNA Restriction Fragments

104EDVO-Kit #



The Exp

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RuNNING ThE GEL

1. After the DNA samples are loaded, carefully snap the cover down onto the electrode terminals. Make sure that the negative and positive color-coded indicators on the cover and apparatus chamber are properly oriented.

2. Insert the plug of the black wire into the black input of the power source (negative input). Insert the plug of the red wire into the red input of the power source (positive input).

3. Set the power source at the required voltage and conduct electrophore-sis for the length of time determined by your instructor.

4. Check to see that current is flowing properly - you should see bubbles forming on the two platinum electrodes.

5. After the electrophoresis is completed, turn off the power, unplug the power source, disconnect the leads and remove the cover.

6. Remove the gel from the bed for staining.

STAINING AND VISuALIzATION OF DNA After electrophoresis, agarose gels require staining to visualize the separated DNA samples. Various options are available for DNA staining. Your instruc-tor will provide instructions for the DNA staining method you will be using.

InstaStain is a registered trademark of EDVOTEK, Inc. Patents Pending.

Electrophoresis can be completed in 15-20 minutes under optimal conditions. For Time and Voltage recommendations, refer to Table C (in Appendix A or B).


14



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Answer the following study questions in your laboratory notebook or on a separate worksheet.

1. How should the x and y axes of the semi-log graph paper used in this experiment, be labeled?

2. Determine unknown 1 and unknown 2 DNA fragment sizes in base pairs according to your standard curve, then determine your percentage error.

3. Use the example of the standard curve to determine:

A. the base pair size if the migration distance is 2.8 centimeters.

B. the migration distance if the fragment contains 5,500 base pairs.

Study Questions

material Safety Data SheetsFull size (8.5 x 11”) pdf copy of MSDS available at www.edvotek.com or by request.

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no

t p

erm

itte

d.

If a

ny

item

is n

ot

app

licab

le, o

r n

o in

form

atio

n is

ava

ilab

le, t

he

spac

e m

ust

b

e m

arke

d t

o in

dic

ate

that

.

Sect

ion

IM

anu

fact

ure

r's

Nam

e

Sect

ion

II -

Haz

ard

ou

s In

gre

die

nts

/Id

enti

fy In

form

atio

n

Emer

gen

cy T

elep

ho

ne

Nu

mb

er

Tele

ph

on

e N

um

ber

fo

r in

form

atio

n

Dat

e Pr

epar

ed

Sig

nat

ure

of

Prep

arer

(o

pti

on

al)

Ad

dre

ss (

Nu

mb

er, S

tree

t, C

ity,

Sta

te,

Zip

Co

de)

EDV

OTE

K, I

nc.

1467

6 R

oth

geb

Dri

veR

ock

ville

, MD

208

50

Haz

ard

ou

s C

om

po

nen

ts [

Spec

ific

C

hem

ical

Iden

tity

; C

om

mo

n N

ame(

s)]

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SHA

PEL

AC

GIH

TLV

Oth

er L

imit

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pti

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(301

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Bo

ilin

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Ph

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hem

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Un

usu

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Spec

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ire

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Pro

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Vap

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or

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ash

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Exti

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uis

hin

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edia

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imit

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ffer

This

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us

mat

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ls a

s d

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ed b

y th

e O

SHA

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mm

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icat

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Sta

nd

ard

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No

dat

a

No

dat

a

No

dat

a

No

dat

a

No

dat

a

No

dat

a

Ap

pre

ciab

le, (

gre

ater

th

an 1

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neg

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Use

ext

ing

uis

hin

g m

edia

ap

pro

pri

ate

for

surr

ou

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ing

fir

e.

Wea

r p

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e eq

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men

t an

d S

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ith

fu

ll fa

cep

iece

op

erat

ed in

po

siti

ve p

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mo

de.

No

ne

iden

tifi

ed

10/0

5/06

Stab

ility

Sect

ion

V -

Rea

ctiv

ity

Dat

aU

nst

able

Sect

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VI -

Hea

lth

Haz

ard

Dat

a

Inco

mp

atib

ility

Co

nd

itio

ns

to A

void

Ro

ute

(s)

of

Entr

y:In

hal

atio

n?

Ing

esti

on

?Sk

in?

Oth

er

Stab

le

Haz

ard

ou

s Po

lym

eriz

atio

nM

ay O

ccu

rC

on

dit

ion

s to

Avo

id

Will

No

t O

ccu

r

Hea

lth

Haz

ard

s (A

cute

an

d C

hro

nic

)

Car

cin

og

enic

ity:

NTP

?O

SHA

Reg

ula

tio

n?

IAR

C M

on

og

rap

hs?

Sig

ns

and

Sym

pto

ms

of

Exp

osu

re

Med

ical

Co

nd

itio

ns

Gen

eral

ly A

gg

rava

ted

by

Exp

osu

re

Emer

gen

cy F

irst

Aid

Pro

ced

ure

s

Sect

ion

VII

- Pr

ecau

tio

ns

for

Safe

Han

dlin

g a

nd

Use

Step

s to

be

Take

n in

cas

e M

ater

ial i

s R

elea

sed

fo

r Sp

illed

Was

te D

isp

osa

l Met

ho

d

Prec

auti

on

s to

be

Take

n in

Han

dlin

g a

nd

Sto

rin

g

Oth

er P

reca

uti

on

s

Sect

ion

VIII

- C

on

tro

l Mea

sure

s

Ven

tila

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nLo

cal E

xhau

stSp

ecia

l

Mec

han

ical

(G

ener

al)

Res

pir

ato

ry P

rote

ctio

n (

Spec

ify

Typ

e)

Pro

tect

ive

Glo

ves

Oth

er P

rote

ctiv

e C

loth

ing

or

Equ

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rk/H

ygie

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ctic

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Eye

Pro

tect

ion

Haz

ard

ou

s D

eco

mp

osi

tio

n o

r B

ypro

du

cts

X

N

on

e

Stro

ng

oxi

diz

ing

ag

ents

Car

bo

n m

on

oxi

de,

Car

bo

n d

ioxi

de

X

N

on

e

Yes

Y

es

Y

es

No

ne

No

ne

iden

tifi

ed

Irri

tati

on

to

up

per

res

pir

ato

ry t

ract

, ski

n, e

yes

No

ne

Ing

esti

on

: If

co

nsc

iou

s, g

ive

larg

e am

ou

nts

of

wat

er

Eyes

: Fl

ush

wit

h w

ater

In

hal

atio

n:

Mo

ve t

o f

resh

air

Sk

in:

Was

h w

ith

so

ap a

nd

wat

er

Wea

r su

itab

le p

rote

ctiv

e cl

oth

ing

. M

op

up

sp

ill

and

rin

se w

ith

wat

er, o

r co

llect

in a

bso

rpti

ve m

ater

ial a

nd

dis

po

se o

f th

e ab

sorp

tive

mat

eria

l.

Dis

po

se in

acc

ord

ance

wit

h a

ll ap

plic

able

fed

eral

, sta

te, a

nd

loca

l en

viro

men

tal r

egu

lati

on

s.

Avo

id e

ye a

nd

ski

n c

on

tact

.

No

ne

Yes

N

on

e

Yes

N

on

e

Yes

_Saf

ety

go

gg

les

No

ne

No

ne

Stab

ility

Sect

ion

V -

Rea

ctiv

ity

Dat

aU

nst

able

Sect

ion

VI -

Hea

lth

Haz

ard

Dat

a

Inco

mp

atib

ility

Co

nd

itio

ns

to A

void

Ro

ute

(s)

of

Entr

y:In

hal

atio

n?

Ing

esti

on

?Sk

in?

Oth

er

Stab

le

Haz

ard

ou

s Po

lym

eriz

atio

nM

ay O

ccu

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on

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Avo

id

Will

No

t O

ccu

r

Hea

lth

Haz

ard

s (A

cute

an

d C

hro

nic

)

Car

cin

og

enic

ity:

NTP

?O

SHA

Reg

ula

tio

n?

IAR

C M

on

og

rap

hs?

Sig

ns

and

Sym

pto

ms

of

Exp

osu

re

Med

ical

Co

nd

itio

ns

Gen

eral

ly A

gg

rava

ted

by

Exp

osu

re

Emer

gen

cy F

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Aid

Pro

ced

ure

s

Sect

ion

VII

- Pr

ecau

tio

ns

for

Safe

Han

dlin

g a

nd

Use

Step

s to

be

Take

n in

cas

e M

ater

ial i

s R

elea

sed

fo

r Sp

illed

Was

te D

isp

osa

l Met

ho

d

Prec

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on

s to

be

Take

n in

Han

dlin

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nd

Sto

rin

g

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s

Sect

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on

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l Mea

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Ven

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ener

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Res

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ry P

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Spec

ify

Typ

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tect

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Oth

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loth

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ard

ou

s D

eco

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osi

tio

n o

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ypro

du

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on

e

Stro

ng

oxi

diz

ing

ag

ents

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bo

n m

on

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ioxi

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nit

rog

en o

xid

es, h

ydro

gen

bro

mid

e g

as

X

N

on

e

Yes

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es

Yes

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dat

a av

aila

ble

Irri

tati

on

to

mu

cou

s m

emb

ran

es a

nd

up

per

res

pir

ato

ry t

ract

No

dat

a

Trea

t sy

mp

tom

atic

ally

an

d s

up

po

rtiv

ely

Wea

r SC

BA

, ru

bb

er b

oo

ts, r

ub

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glo

ves

Mix

mat

eria

l wit

h c

om

bu

stib

le s

olv

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and

bu

rn in

a c

hem

ical

inci

ner

ato

r eq

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ped

aft

erb

urn

er a

nd

scr

ub

ber

Use

in c

hem

ical

fu

me

ho

od

wit

h p

rop

er p

rote

ctiv

e la

b g

ear.

Mu

tag

en

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em. f

um

e h

oo

d

No

N

on

e

Ru

bb

er

C

hem

. saf

ety

go

gg

les

R

ub

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bo

ots

Use

in c

hem

ical

fu

me

ho

od

wit

h p

rop

er p

rote

ctiv

e la

b g

ear.

Acu

te: M

ater

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rrit

atin

g t

o m

uco

us

mem

bra

nes

, up

per

res

pir

ato

ry t

ract

, eye

s, s

kin

Ch

ron

ic:

May

alt

er g

enet

ic m

ater

ial

SCB

A

Mat

eria

l Saf

ety

Dat

a Sh

eet

May

be

use

d t

o c

om

ply

wit

h O

SHA

's H

azar

d C

om

mu

nic

atio

nSt

and

ard

. 29

CFR

191

0.12

00 S

tan

dar

d m

ust

be

con

sult

ed f

or

spec

ific

req

uir

emen

ts.

IDEN

TITY

(A

s U

sed

on

Lab

el a

nd

Lis

t)N

ote

: B

lan

k sp

aces

are

no

t p

erm

itte

d.

If a

ny

item

is n

ot

app

licab

le, o

r n

o in

form

atio

n is

ava

ilab

le, t

he

spac

e m

ust

b

e m

arke

d t

o in

dic

ate

that

.

Sect

ion

IM

anu

fact

ure

r's

Nam

e

Sect

ion

II -

Haz

ard

ou

s In

gre

die

nts

/Id

enti

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form

atio

n

Emer

gen

cy T

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ne

Nu

mb

er

Tele

ph

on

e N

um

ber

fo

r in

form

atio

n

Dat

e Pr

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ed

Sig

nat

ure

of

Prep

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pti

on

al)

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in, I

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P.O

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x 12

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est

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hes

da,

MD

208

27

Haz

ard

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po

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ts [

Spec

ific

C

hem

ical

Iden

tity

; C

om

mo

n N

ame(

s)]

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SHA

PEL

AC

GIH

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) 25

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Ph

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Ch

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Un

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edia

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Eth

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m B

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Eth

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D

ata

no

t av

aila

ble

No

dat

a

No

dat

a

No

dat

a

No

dat

a

No

dat

a

No

dat

a

Solu

ble

Ch

emic

al b

ou

nd

to

pap

er, n

o o

do

r

No

dat

a

N.D

. = N

o d

ata N.D

. N

.D.

Wat

er s

pra

y, c

arb

on

dio

xid

e, d

ry c

hem

ical

po

wd

er, a

lco

ho

l or

po

lym

er f

oam

Wea

r p

rote

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e cl

oth

ing

an

d S

CB

A t

o p

reve

nt

con

tact

wit

h s

kin

& e

yes

Emit

s to

xic

fum

es

10/0

5/06

CA

S# 1

39-3

3-3

(2,7

-Dia

min

o-1

0-Et

hyl

-9-P

hen

ylp

hen

anth

rid

iniu

m B

rom

ide)

ED

VO

TE

K®

material Safety Data SheetsFull size (8.5 x 11”) pdf copy of MSDS available at www.edvotek.com or by request.

Mat

eria

l Saf

ety

Dat

a Sh

eet

May

be

use

d t

o c

om

ply

wit

h O

SHA

's H

azar

d C

om

mu

nic

atio

nSt

and

ard

. 29

CFR

191

0.12

00 S

tan

dar

d m

ust

be

con

sult

ed f

or

spec

ific

req

uir

emen

ts.

IDEN

TITY

(A

s U

sed

on

Lab

el a

nd

Lis

t)N

ote

: B

lan

k sp

aces

are

no

t p

erm

itte

d.

If a

ny

item

is n

ot

app

licab

le, o

r n

o in

form

atio

n is

ava

ilab

le, t

he

spac

e m

ust

b

e m

arke

d t

o in

dic

ate

that

.

Sect

ion

IM

anu

fact

ure

r's

Nam

e

Sect

ion

II -

Haz

ard

ou

s In

gre

die

nts

/Iden

tify

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rmat

ion

Emer

gen

cy T

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ne

Nu

mb

er

Tele

ph

on

e N

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ber

fo

r in

form

atio

n

Dat

e Pr

epar

ed

Sig

nat

ure

of

Prep

arer

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pti

on

al)

Ad

dre

ss (

Nu

mb

er, S

tree

t, C

ity,

Sta

te,

Zip

Co

de)

EDV

OTE

K, I

nc.

1467

6 R

oth

geb

Dri

veR

ock

ville

, MD

208

50

Haz

ard

ou

s C

om

po

nen

ts [

Spec

ific

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hem

ical

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tity

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om

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SHA

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ash

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Exti

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edia

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mab

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imit

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n R

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ED

VO

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K®

Inst

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Met

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Phen

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n 5

IUM

C

hlo

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No

dat

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CA

S #

61-7

3-4

No

dat

a

No

dat

a

No

dat

a

No

dat

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No

dat

a

No

dat

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Solu

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old

Ch

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dat

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N

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Wat

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Emit

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fu

mes

un

der

fir

e co

nd

itio

ns

Stab

ility

Sect

ion

V -

Rea

ctiv

ity

Dat

aU

nst

able

Sect

ion

VI -

Hea

lth

Haz

ard

Dat

a

Inco

mp

atib

ility

Co

nd

itio

ns

to A

void

Ro

ute

(s)

of

Entr

y:In

hal

atio

n?

Ing

esti

on

?Sk

in?

Oth

er

Stab

le

Haz

ard

ou

s Po

lym

eriz

atio

nM

ay O

ccu

rC

on

dit

ion

s to

Avo

id

Will

No

t O

ccu

r

Hea

lth

Haz

ard

s (A

cute

an

d C

hro

nic

)

Car

cin

og

enic

ity:

NTP

?O

SHA

Reg

ula

tio

n?

IAR

C M

on

og

rap

hs?

Sig

ns

and

Sym

pto

ms

of

Exp

osu

re

Med

ical

Co

nd

itio

ns

Gen

eral

ly A

gg

rava

ted

by

Exp

osu

re

Emer

gen

cy F

irst

Aid

Pro

ced

ure

s

Sect

ion

VII

- Pr

ecau

tio

ns

for

Safe

Han

dlin

g a

nd

Use

Step

s to

be

Take

n in

cas

e M

ater

ial i

s R

elea

sed

fo

r Sp

illed

Was

te D

isp

osa

l Met

ho

d

Prec

auti

on

s to

be

Take

n in

Han

dlin

g a

nd

Sto

rin

g

Oth

er P

reca

uti

on

s

Sect

ion

VIII

- C

on

tro

l Mea

sure

s

Ven

tila

tio

nLo

cal E

xhau

stSp

ecia

l

Mec

han

ical

(G

ener

al)

Res

pir

ato

ry P

rote

ctio

n (

Spec

ify

Typ

e)

Pro

tect

ive

Glo

ves

Oth

er P

rote

ctiv

e C

loth

ing

or

Equ

ipm

ent

Wo

rk/H

ygie

nic

Pra

ctic

es

Eye

Pro

tect

ion

Haz

ard

ou

s D

eco

mp

osi

tio

n o

r B

ypro

du

cts

X

No

ne

Stro

ng

oxi

diz

ing

ag

ents

Toxi

c fu

mes

of

Car

bo

n m

on

oxi

de,

Car

bo

n d

ioxi

de,

n

itro

gen

oxi

des

, su

lfu

r o

xid

es, h

ydro

gen

, ch

lori

de

gas

X

N

on

e

Yes

Y

es

Yes

Skin

: M

ay c

ause

ski

n ir

rita

tio

n

Eyes

: M

ay c

ause

eye

irri

tati

on

In

hal

atio

n:

Cya

no

sis

Mee

ts c

rite

ria

for

pro

po

sed

OSH

A m

edic

al r

eco

rds

rule

PER

EAC

47.

3042

0.82

No

dat

a av

aila

ble

No

dat

a av

aila

ble

Trea

t sy

mp

tom

atic

ally

Ven

tila

te a

rea

and

was

h s

pill

sit

e

Mix

mat

eria

l wit

h a

co

mb

ust

ible

so

lven

t an

d b

urn

in c

hem

ical

inci

ner

ato

r eq

uip

ped

wit

h a

fter

bu

rner

an

d s

cru

bb

er.

Ch

eck

loca

l an

d s

tate

reg

ula

tio

ns.

Kee

p t

igh

tly

clo

sed

. St

ore

in c

oo

l, d

ry p

lace

No

ne

MIO

SH/O

SHA

ap

pro

ved

, SC

BA

Req

uir

ed

Ru

bb

erC

hem

. saf

ety

go

gg

les

Ru

bb

er b

oo

ts

Stab

ility

Sect

ion

V -

Rea

ctiv

ity

Dat

aU

nst

able

Sect

ion

VI -

Hea

lth

Haz

ard

Dat

a

Inco

mp

atib

ility

Co

nd

itio

ns

to A

void

Ro

ute

(s)

of

Entr

y:In

hal

atio

n?

Ing

esti

on

?Sk

in?

Oth

er

Stab

le

Haz

ard

ou

s Po

lym

eriz

atio

nM

ay O

ccu

rC

on

dit

ion

s to

Avo

id

Will

No

t O

ccu

r

Hea

lth

Haz

ard

s (A

cute

an

d C

hro

nic

)

Car

cin

og

enic

ity:

NTP

?O

SHA

Reg

ula

tio

n?

IAR

C M

on

og

rap

hs?

Sig

ns

and

Sym

pto

ms

of

Exp

osu

re

Med

ical

Co

nd

itio

ns

Gen

eral

ly A

gg

rava

ted

by

Exp

osu

re

Emer

gen

cy F

irst

Aid

Pro

ced

ure

s

Sect

ion

VII

- Pr

ecau

tio

ns

for

Safe

Han

dlin

g a

nd

Use

Step

s to

be

Take

n in

cas

e M

ater

ial i

s R

elea

sed

fo

r Sp

illed

Was

te D

isp

osa

l Met

ho

d

Prec

auti

on

s to

be

Take

n in

Han

dlin

g a

nd

Sto

rin

g

Oth

er P

reca

uti

on

s

Sect

ion

VIII

- C

on

tro

l Mea

sure

s

Ven

tila

tio

nLo

cal E

xhau

stSp

ecia

l

Mec

han

ical

(G

ener

al)

Res

pir

ato

ry P

rote

ctio

n (

Spec

ify

Typ

e)

Pro

tect

ive

Glo

ves

Oth

er P

rote

ctiv

e C

loth

ing

or

Equ

ipm

ent

Wo

rk/H

ygie

nic

Pra

ctic

es

Eye

Pro

tect

ion

Haz

ard

ou

s D

eco

mp

osi

tio

n o

r B

ypro

du

cts

X

N

on

e

No

ne

Sulf

ur

oxi

des

, an

d b

rom

ides

X

N

on

e

Yes

Y

es

Yes

Acu

te e

ye c

on

tact

: M

ay c

ause

irri

tati

on

. N

o d

ata

avai

lab

le f

or

oth

er r

ou

tes.

No

dat

a av

aila

ble

May

cau

se s

kin

or

eye

irri

tati

on

No

ne

rep

ort

ed

Trea

t sy

mp

tom

atic

ally

an

d s

up

po

rtiv

ely.

Rin

se c

on

tact

ed a

rea

wit

h c

op

iou

s am

ou

nts

of

wat

er.

Wea

r ey

e an

d s

kin

pro

tect

ion

an

d m

op

sp

ill a

rea.

Rin

se w

ith

wat

er.

Ob

serv

e al

l fed

eral

, sta

te, a

nd

loca

l reg

ula

tio

ns.

Avo

id e

ye a

nd

ski

n c

on

tact

.

No

ne

Yes

No

ne

Yes

No

ne

Yes

Spla

sh p

roo

f g

og

gle

s

No

ne

req

uir

ed

Avo

id e

ye a

nd

ski

n c

on

tact

Mat

eria

l Saf

ety

Dat

a Sh

eet

May

be

use

d t

o c

om

ply

wit

h O

SHA

's H

azar

d C

om

mu

nic

atio

nSt

and

ard

. 29

CFR

191

0.12

00 S

tan

dar

d m

ust

be

con

sult

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or

spec

ific

req

uir

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ts.

IDEN

TITY

(A

s U

sed

on

Lab

el a

nd

Lis

t)N

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k sp

aces

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itte

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If a

ny

item

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app

licab

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r n

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form

atio

n is

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he

spac

e m

ust

b

e m

arke

d t

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dic

ate

that

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Sect

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IM

anu

fact

ure

r's

Nam

e

Sect

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II -

Haz

ard

ou

s In

gre

die

nts

/Id

enti

fy In

form

atio

n

Emer

gen

cy T

elep

ho

ne

Nu

mb

er

Tele

ph

on

e N

um

ber

fo

r in

form

atio

n

Dat

e Pr

epar

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Sig

nat

ure

of

Prep

arer

(o

pti

on

al)

Ad

dre

ss (

Nu

mb

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tree

t, C

ity,

Sta

te,

Zip

Co

de)

EDV

OTE

K, I

nc.

1467

6 R

oth

geb

Dri

veR

ock

ville

, MD

208

50

Haz

ard

ou

s C

om

po

nen

ts [

Spec

ific

C

hem

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Iden

tity

; C

om

mo

n N

ame(

s)]

O

SHA

PEL

AC

GIH

TLV

Oth

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imit

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eco

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pti

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(301

) 25

1-59

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(301

) 25

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Bo

ilin

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oin

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III -

Ph

ysic

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hem

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Ch

arac

teri

stic

s

Un

usu

al F

ire

and

Exp

losi

on

Haz

ard

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Spec

ial F

ire

Fig

hti

ng

Pro

ced

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or

Pres

sure

(m

m H

g.)

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or

Den

sity

(A

IR =

1)

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bili

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Wat

er

Ap

pea

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or

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ion

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ysic

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int

(Met

ho

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Exti

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hin

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edia

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mab

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imit

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ELLE

L

Mel

tin

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oin

t

Evap

ora

tio

n R

ate

(Bu

tyl A

ceta

te =

1)

Spec

ific

Gra

vity

(H

0 =

1)

2

Prac

tice

Gel

Lo

adin

g S

olu

tio

n

10/0

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pro

du

ct c

on

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o h

azar

do

us

mat

eria

ls a

s d

efin

ed b

y th

e O

SHA

Haz

ard

Co

mm

un

icat

ion

Stan

dar

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No

dat

a

No

dat

a

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dat

a

No

dat

a

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dat

a

No

dat

a

Solu

ble

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lue

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o o

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o d

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dat

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ch

emic

al, c

arb

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ater

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Use

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ear

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A.

Un

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ED

VO

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The Biotechnology Education Company · The Biotechnology Education Company ® EDVOTEK, Inc. •...

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Transcript of The Biotechnology Education Company · The Biotechnology Education Company ® EDVOTEK, Inc. •...