BD_Reagents_Apoptosis_Brochure.pdf

7/27/2019 BD_Reagents_Apoptosis_Brochure.pdf

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Apoptosis, Cell Cycle, and

Cell Prolieration

Tools for Assessing Cell Events


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3/163For Research Use Only. Not for use in diagnostic or therapeutic procedures.

Life, Death, and Cell ProliferationThe balance o cell prolieration and apoptosis is important or both development and normal

tissue homeostasis. Cell prolieration is an increase in the number o cells as a result o growth

and division. Cell prolieration is regulated by the cell cycle, which is divided into a series o

phases. Apoptosis, or programmed cell death, results in controlled sel-destruction.

Several methods have been developed to assess apoptosis, cell cycle, and cell prolieration.

BD Biosciences oers a complete portolio o reagents and tools to allow exploration o the

cellular eatures o these processes.

Over the years multicolor low cytometry has become essential in the study o apoptosis,

cell cycle, and cell prolieration. Success o the technology results rom its ability to monitor

these processes along with other cellular events, such as protein phosphorylation or cytokine

secretion, within heterogeneous cell populations. BD Biosciences continues to innovate in

this area with new products such as the BD Horizon violet cell prolieration dye 450

(VPD450) and popular reagents such as antibodies to cleaved PARP and caspase-3 available

in new ormats and or dierent types o applications.

In addition to low cytometry products, BD Biosciences carries a broad portolio o

reagents or determination and detection o apoptotic and prolierative events by ELISA,

immunohistochemistry, cell imaging, or Western blot.

As part o our commitment to maximize scientiic results, BD Biosciences provides a variety

o tools to assist customers in their experimental setup and analysis. These include a decisiontree to guide in the selection o the most suitable methods or a speciic study.

BD Biosciences carries high-quality reagents in the latestformats to examine cell cycle, proliferation, and apoptosisacross a variety of platforms, in applications from basicresearch to drug screening.


4/164 For Research Use Only. Not for use in diagnostic or therapeutic procedur

Fundamental cellular processes

Cell Cycle and Cell Proliferation:An Overview

To help researchers better understand the undamental

cellular mechanisms involved in immunity, inlammation,

hematopoiesis, neoplasia, and other biological responses,

BD Biosciences oers a range o tools including antibodies,

kits, and systems to measure prolierative responses. Using

low cytometry, immunoluorescence, or immunohisto-

chemistry, researchers can quickly and accurately determine

the cell cycle status or tissue localization o individual cellswithin prolierating populations. These tools include:

BD Biosciences reagents and BD Cycletest Plus reagent

kit or the analysis o cellular DNA content

DNA dyes, propidium iodide (PI), 7-aminoactinomycin D

(7-AAD)

Antibodies against cyclins, retinoblastoma, and

phosphorylated histone H3

In adaptive immunity, speciic T and B lymphocytes undergo

clonal expansion (division, prolieration, and dierentiation)

in response to oreign antigenic stimulation. Cell growth,

replication, and division in eukaryotic cells occur accordingto a highly controlled series o events called the cell cycle.1

The Cell Cycle

The cell cycle has two major phases: interphase, the phase

between mitotic events, and the mitotic phase, where the

mother cell divides into two genetically identical daughte

cells. Interphase has three distinct, successive stages.

During the irst stage called G1, cells monitor their

environment, and when the requisite signals are received,

the cells synthesize RNA and proteins to induce growth.When conditions are right, cells enter the S stage o the c

cycle and commit to DNA synthesis and replicate their

chromosomal DNA. Finally in the G2 phase cells continue t

grow and prepare or mitosis.

Cell cycle phase

Interphase

Preparationfor

Mitosis Preparationfor DNA

SynthesisGrowth

Telo

phase

Prophase

Metap

hase

Anaphase

Growth

DNAReplication

G2 G1

G0

M

S

Measures Reagents Mechanism Technology Sample Types

DNA Propidium Iodide (PI), 7-aminoactino-mycin D (7-AAD)

Interaction into DNA double strands Flow cytometry Fixed, permeabilized, and or live/deaddiscrimination in intact cells

Cell Dyes BD Horizon violet prolieration dye 450

(VPD450)

Diuses into live cells and is hydrolyzed

by intracellular non-specic esterasesto become fuorescent products.

Flow cytometry Live prolierating cells

Newly Synthesized DNA BrdU and antibodies to BrdU Bromodeoxy uridine replaces thymidine(T) in dividing DNA. It is then detectedby antibodies to BrdU.

Flow cytometry, cell imaging,immunohistochemistry

Fixed and permeabilized cells, treatedtissues (cell imaging, immunohisto-chemistry only)

Protein Level Antibodies to Ki67, PCNA Levels increase as a result oprolieration.

Flow cytometry, bioimaging,immunohistochemistry, Western blot

Fixed cells, tissues, and extracts

Protein Level Antibodies to cyclins, retinoblastoma(Rb), other cell cycle markers

Levels go up and down at dierentstages o the cell cycle.


Fixed cells, tissues, and extracts

Protein Modication Antibodies to phosphorylated histoneH3, cyclin dependent kinases (cdk)

Proteins become phosphorylated asa result o prolieration or changes tothe cell cycle.


BD CBA (or quantitative detection) Fixed cells, tissues, and extracts

Methods or the study o cell cycle and prolieration



Analysis of Cellular DNA Content

BD Biosciences oers a wide variety o reagents to study

the cell cycle. Reagents include DNA dyes such as propidium

iodide (PI) and 7-amino actinomycin D (7-AAD). In addition,

the BD Cycletest Plus reagent kit includes PI and other

reagents to degrade proteins and RNA to allow more

precise DNA measurement. The samples are subsequently

analyzed using low cytometry to assess ploidy, identiyabnormal DNA stemlines, and estimate the DNA index (DI)

and cell cycle phase distributions o stemlines.

During the cell cycle phases, DNA levels change, acilitating

the use o DNA dyes such as 7-AAD to generate characteristic

cellular DNA content proiles (see the igure below).

As cells go through the phases o the cell cycle, proteins

such as histone H3 Ser28 become modiied or change

in expression.2 To acilitate DNA replication the histone

is modiied, opening the chromatin to allow entry o

replication machinery. To urther support the study o cell

cycle, BD Biosciences carries antibodies to these proteins to

use or imaging or low cytometry applications.

CD4 enriched mouse splenocytes were

cultured with anti-CD3/CD28, IL-2, and IL-4

or 6 days. Cells were harvested and treated

with 10 ng/mL IL-2+1 g/mL colcemid or 4

(top) or 24 (bottom) hours and stained with

the BD Cycletest Plus DNA reagent kit.

Count

PE-A

50 100 150 200 250

250

500

750

1,

000

(x 1,000)

BrdU(

APC)

7-AAD (FL3)

50 100 150 200 250

101

102

103

104

R4

R5R3

R6BrdU(

FITC)

7-AAD (FL3)

50 100 150 200 250

101

102

103

104

R4

R5

R3

R6

Count

PE-A

50 100 150 200 250

50

100

150

200

250

300

(x 1,000)

Cell cycle analysis of a populationstained for incorporated BrdU and

total DNA levels (7-AAD).

Human PBMCs were stimulated with

anti-CD3/CD28 or 48 hours and

re-stimulated with PMA+ionomycin

or 4 hours, and BrdU was added

or the inal 1 hour. Cells were then

harvested and stained using the

BrdU staining protocol.



New tool to determine cell divisions

Tools and Techniques to StudyCell Proliferation

Cell prolieration can occur in response to many stimuli

such as cytokine exposure or a variety o other processes.

BD has a new product to help researchers study cell

prolieration.

BD Biosciences oers Violet Prolieration Dye 450 or

the detection o cell prolieration with the violet laser,

which acilitates the use o larger panels. This allows the

determination o more data rom limited samples usingmulticolor low cytometry.

VPD450 is a nonluorescent esteriied dye. The ester group

allows the dye to enter the cell. Once the dye is inside the

cell, esterases cleave o the ester group to convert the

dye into a luorescent product and trap it inside the cell.

With each replication event the amount o dye in the cell

decreased, leading to a characteristic pattern.

The use of VPD450 to correlate cell proliferation with IL-2production.

CD4+ enriched mouse splenocytes were loaded with 1 M VPD450

or 10 minutes. Cells were then stimulated with anti-CD3/CD28

and harvested at the indicated times. Approximately 4 to 6 hours

prior to harvest, cells were stimulated with PMA/ionomycin in the

presence o BD GolgiStop protein transport inhibitor. Cells were

ixed and permeabilized, stained or IL-2, and analyzed on a

BD LSR II low cytometer.

IL-2AlexaFluor4

88

VPD450

102 103 104 105

102

103

104

105

IL-2AlexaFluor4

88

VPD450

102 103 104 105

102

103

104

105

IL-2AlexaFluor4

88

VPD450

102

103

104

105

102

103

104

105

Day 1 Day 2 Day 3



Tools for BrdU Analysis

BD Biosciences carries a series o antibodies and kits

designed or the detection o prolierating cells by

measurement o bromodeoxyuridine (BrdU), an analog

o the DNA precursor thymidine used to measure de novo

DNA synthesis. During the S phase o the cell cycle (DNA

synthesis) BrdU is incorporated into the newly synthesized

DNA and can be readily detected by anti-BrdU speciicantibodies. BD antibodies and kits designed or the

detection o BrdU are available or both intracellular

low cytometry and immunohistochemistry and include

BD Horizon V450 and PerCP-Cy 5.5 ormats.

In addition to DNA increases, levels o certain proteins also

rise as a result o cell prolieration. For example, Ki67 is an

antigen that is expressed in the nucleus o dividing cells.

However, during the G0 phase o the cell cycle it is not

detected. Ki67 can be combined with other prolieration

markers such as BrdU and VPD450 or added conidence.

These markers can also be combined with cell surace and

other types o markers to gain additional inormation aboutcell subsets and their signaling pathways.

Cell cycle analysis o HeLa cells treated with Aphidicolin (DNApolymerase inhibitor) monitored by BrdU staining. The images

were captured on a BD Pathway 855 bioimaging system with

a 20x objective and merged using BD Attovision sotware.

Hoechst Blue

BrdU Red

Histone H3 (pS28) Yellow

Tubulin Green

Untreated

1g Aphidicolin



The importance o tissue homeostasis

As cells become damaged or are no longer needed, they

undergo apoptosis or programmed cell death, a normal

physiological process that occurs during embryonic

development and tissue homeostasis maintenance.

Apoptosis is an organized process that signals cells to

sel destruct or cell renewal or to control aberrant cell

growth. Apoptosis controls the orderly death o damaged

cells, whereas necrosis occurs as a result o tissue damage,causing the loss o both damaged and surrounding cells.3

The apoptotic process is characterized by certain morpho-

logical eatures. These include changes in the plasma

membrane (such as loss o membrane symmetry and loss

o membrane attachment), a condensation o the cytoplasm

and nucleus, protein cleavage, and internucleosomal

cleavage o DNA. In the inal stages o the process, dying

cells become ragmented into apoptotic bodies and

consequently are eliminated by phagocytic cells without

signiicant inlammatory damage to surrounding cells.

However, some cell types do not display characteristic

eatures o apoptosis. In those cases multiple aspects

o apoptosis might need to be analyzed to conirm the

mechanism o cell death.

To support this spectrum o requirements, BD Biosciences

oers a ull range o apoptosis detection tools and

technologies or measuring indicators at dierent stages

across the apoptotic process. BD Biosciences tools use multipmethodologies including low cytometry, bioimaging, and

microscopy (or live and ixed cell analysis) as well as ELISA

IHC, Western blot, and spectroluorometry.

Techniques to Study ApoptosisProgrammed Cell Death

Intact Cells

Flow Cytometry Bioimaging/Microscopy

Live Fixed

IHC

Cell Extracts Tissue Sections

Apoptotic Cells

Loss of Membrane

Asymmetry Annexin V

Cleaved Markers

Active Caspase-3 Cleaved PARP

MitochondrialMembrane Potential

MitoScreen (JC-1)

TUNEL/DNAFragmentation

APO-DIRECT APO-BRDU

Live Fixed

Western Blot or

Immunoprecipitation

Annexin V Active Caspase-3

Cleaved PARP

Apoptosis Markers

Spectrofluorometry

Caspase Activity

Active Caspase-3 Cleaved PARP Active Caspase-3 Cleaved PAR

ELISA


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For Research Use Only. Not for use in diagnostic or therapeutic procedures.

Annexin VA Key Protein in Apoptosis Signaling

Changes in the plasma membrane are one o the irst

characteristics o the apoptotic process detected in living

cells. Apoptosis can be detected by the presence ophosphatidylserine (PS), which is normally located on the

cytoplasmic ace o the plasma membrane. During apoptosis

PS translocates to the outer lealet o the plasma membrane

and can be detected by low cytometry and cell imaging

through binding to luorochrome-labeled Annexin V when

calcium is present.

BD Biosciences oers Annexin V in several common ormats

such as FITC, PE, and BD Horizon V450 or the violet laser.

With the addition o these new ormats, more complex

assays can be developed to look at apoptosis withinheterogeneous cell subsets.

Since intracellular Annexin V is also exposed i the plasma

membrane is compromised, a membrane-impermeant dye

such as 7-AAD is commonly used to distinguish between

apoptotic and dead cells to exclude the dead cells. The

populations o cells that are stained with Annexin V only

represent the apoptotic cell populations.

Ca++

Annexin V-PE Conjugate

Normal CellCytoplasm

Plasma Membrane Phospholipid Flipping

Externalization ofPhosphatidylserine

Apoptosis

Apoptotic CellCytoplasm

Ca++ Ca++

Radio requency dose dependent apoptosis, necrosis, and cell deathmonitored by Annexin V-BD Horizon V450 in pancreatic carcinoma

cell lines treated with a low dose o cetuximab targeted gold

nanoparticles. As the RF ield power increases, the temperature

increases, and a shit rom apoptosis (lower right quadrant) to

rank necrosis (upper let quadrant) is seen.

Data courtesy o ES Glazer and SA Curley, MD Anderson Cancer Center.

7-AAD

Annexin V - BD Horizon V450

102 103 104 1050

102

103

104

105

0

Low RF Field Treatment

7-AAD


102 103 104 1050

102

103

104

105

0

High RF Field Treatment

7-AAD


Control

102 103 104 1050

102

103

104

105

0



Extracellular SpaceTNF/FasL

Growth FactorsCytokines

RTKTNFR/Fas

Caspace8/10

DNA DamageASK1

Ras

MKK4/7

c-Raf

PLCY

MEK1/2

ERK1/2 p90RSK

Cdc2BADBcl-2Mitochondria

BAK

Bfl-1

tBid

BAXBcl-XL

Endo G

AIF

CytochromeC

Diablo

HtrA2

HtrA2

Diablo CytochromeC

AIF Endo G

Cytoplasm

tBid

Bid

Caspace2 p53 JNK1

cIAP

Apaf1

Caspace9

Caspace6

Caspace3

CAD ICAD

ROCK1 Gas2 FodrinLamin

A

Cell Shrinkage &Membrane Blebbing

Caspace-independentDNA Fragmentation

DNAFragmentation

Apoptosis

ChromatinCondensation

DNARepair

PARP Acinus

Caspace7

PKC(,,)

There are many apoptosis triggers including certain

cytokines, protein-protein interactions, and chemicals. Once

apoptosis starts, changes in the mitochondria membrane

potential can be measured by low cytometry using the

BD MitoScreen (JC-1) low cytometry kit.

Increases in mitochondrial membrane potential lead to

increased mitochondrial membrane permeability and the

release o soluble proteins such as cytochrome c and

pro-caspases.

Caspases are a series o proteases activated upon cleavage

at aspartate residues during earliest stages o apoptosis.

Active caspases can then cleave many proteins including

Poly-ADP ribose polymerase (PARP) and other caspases.

DNA ragmentation is one o the last phases in apoptosis

resulting rom the activation o endonucleases during the

apoptotic process. There are several established methods

or the study o DNA ragmentation including isolation an

separation o DNA ragments by agarose gel electrophore

and end labeling.

The BD APO-BrdU kit uses end labeling or the terminal

deoxynucleotidyl transerase (TdT) nick end labeling (TUNE

method) to support the study o DNA ragmentation. In

this assay, TdT catalyzes a template-independent addition

o bromolated deoxyuridine triphosphates (Br-dUTP) to the

3-hydroxyl (OH) termini o double- and single-stranded

DNA. Ater the Br-dUTP is incorporated, these terminal site

o double- and single-stranded DNA are identiied using

low cytometry by staining cells with labeled anti-BrdU. In

contrast, the BrdU prolieration assay incorporates BrdU in

newly synthesized DNA, into sites o DNA strand breaks.

Tools to streamline apoptosis research

Additional Techniques for theDetection of Apoptosis

With an overwhelming number o available techniques and products, selecting the most appropriate

method is oten diicult. To help make this choice easier, the overview above summarizes commercially

available assays rom a biological perspective.

BD Cell Pathways tool

To support signaling research, the BD

Biosciences website includes the BD Cell

Pathways tool, powered by Ingenuity

Systems, to help researchers explore the

pathways that involve target molecules

o interest.

Feature Measured Assays Key FeaturesPlasma Membrane Alterations(Phosphatidylserine Exposure)

Annexin binding assay Single conjugates Annexin V kits

Detects early apoptosis markers Quick and easy Flow cytometry or immunofuorescence application

Mitochondrial Changes BD MitoScreen Kit Fast, easy, single cell resolution by fow cytometry or fuorescentmicroscopy

Caspase Activation Caspase Activity Assay Kits and Reagents

Active Caspase-3 immunoassays

Quick and easy, uses spectrofuorometry

ELISA, fow cytometry, or Western blot

DNA Fragmentation APO-BrdU TUNEL Assay APO-DIRECT TUNEL Assay

Works with adherent cells, single cell resolution in conjunctionwith cell cycle analysis by fow cytometry



Measurement of Cleaved Caspases and PARP

Caspases are important initiators o apoptosis. One o the

earliest and most consistently observed characteristics o

apoptosis is the activation o a series o cytosolic proteases,

called caspases. When apoptosis is activated, caspases cleave

multiple protein substrates en masse, which leads to the

loss o cellular structure and unction, and ultimately results

in cell death.4

In particular, caspases -8, -9, and -3 have beenimplicated in apoptosis: caspase-9 in the mitochondrial

pathway, caspase-8 in the Fas/CD95 pathway, and caspase-3

more downstream, activated by multiple pathways.

BD Biosciences carries a variety o reagents to measure

caspases, particularly caspase-3. They include antibodies

directed exclusively against the active orm o the caspase.

These antibodies are available in a variety o ormats and can

be used or low cytometry, imaging, ELISA, and Western blot.

BD Biosciences oers a range o tools or caspase activity

assays rom individual luorogenic peptide substrates and

inhibitors, to kits, to ready-to-use assay plates. All are based

on the use o synthetic tetrapeptide substrates5 that aredesigned such that proteolytic cleavage by active human

or mouse caspases results in release o a luorophore

or chromophore. The individual synthetic tetrapeptide

substrates, together with the caspase inhibitors and active

caspase enzymes, oer lexibility in the experimental

design o a caspase activity assay.

Caspase-3 cleavage/inhibition reactions

Active caspase-3 binds to the luorogenic Ac-DEVD-AMC

substrate and cleaves it between asparatic acid (D) andAMC, releasing the luorescent AMC. AMC luorescence is

quantiied by UV spectroluorometry. The Ac-DEVD-CHO

aldehyde inhibitor binds strongly to the caspase-3 active

site and blocks substrate binding. Hence, Ac-DEVD-AMC is

not cleaved and luorescence is not emitted.

Flow cytometric analysis of apoptotic and non-apoptoticpopulations using anti-active caspase-3 antibodies.

Jurkat T cells (A, A1) or mouse thymocytes (B, B1) were let

untreated (A, B) or treated or 4 h with camptothecin (A1)

or a mouse Fas monoclonal antibody, clone Jo2 (Cat. No.

554254) to induce apoptosis (B1). Cells were permeabilized

and then stained with PE-conjugated active caspase-3

antibodies (Cat. No. 557091). Untreated cells were primarily

negative or the presence o active caspase-3, whereas

about hal o each population o cells induced to undergo

apoptosis had detectable active caspase-3.

Re

lativeCellNumber

101 102 103

4

0

80

120

160

200

Untreated

95%

A

M15%

M2

RelativeCellNumber

101

102

103

40

80

120

160

200

Treated with Camptothecin (4 h)

48%

A1

M152%

M2

10

3

101

102

103

60

120

180

240

300

Treated with mouse Fas mAb, clone Jo2 (6 h)

55%

B1

M1 45%

M2

101 102 103

60

120

180

240

300

Untreated

99%

B

M1 1%

M2

Active Caspase-3

Cleavage

DEVD-AMC DEVD + AMC

Active Caspase-3

Inhibition

DEVD-CHO DEVD-CHO + DEVD-AMC (No signal)



Obtain the complete picture

Additional Proteins for theStudy of Apoptosis

In addition to caspases and Annexin V, there are several other

proteins important or the study o apoptosis, including the

Bcl-2 amily, tumor necrosis actor receptor (TNFR) amily,

PARP, and other signaling molecules. Bcl-2 amily members,

identiied by the presence o conserved BCL2 homology (BH3)

domains, are versatile key regulators o apoptosis. Bcl-2, or

example, protects cells rom apoptosis by associating with

the mitochondrial membrane and preventing the release ocytochrome c rom the mitochondria. In contrast other Bcl-2

amily members such as Bax promote apoptosis. Increased

levels o Bcl-2 have been reported in cancer.6

In this experiment, Jurkat cells were treated with camptothecin,

a potent inhibitor o topoisomerase I and apoptosis inducer.

Phosphorylation o H2AX, a protein important or maintaining

genome integrity, has been shown to correlate with levels o

DNA damage.10 Using multicolor low cytometry, cell prolieration

(BrdU), apoptosis (cleaved PARP), and DNA damage (histone H2AX

pS140) were evaluated in the same experiment.

The TNFR amily contains many members, including CD95,

that can be divided into three major groups based on

structure. Signaling through the TNFR pathway leads to

apoptosis.7

PARPs are DNA repair enzymes that are activated by DNA

strand breaks. Cleavage o PARP by caspase-3 into 24- and

89-kDa ragments inactivates the PARP enzyme.

BD Biosciences carries antibodies speciic or cleavage

products o PARP that are useul markers o apoptosis.

These antibodies are available in a variety o ormats and

can be combined with other markers to gain additional

inormation about the cell.8,9

PhosphoH2AX-AlexaFlu

or6

47

BrdU - FITC

102 103 104 105

102

103

104

105

Untreated

3% 24%

59% 14%

102 103 104 105

102

103

104

105

20 M Camptothecin

19% 30%

43% 7%

102 103 104 105

102

103

104

105

10 M Camptothecin

20% 29%

44% 7%

102 103 104 105

102

103

104

105

5 M Camptothecin

12% 12%

51% 6%

102 103 104 105

102

103

104

105

2.5 M Camptothecin

7% 30%

56% 7%



Simultaneous Studies of Apoptosis, Cell Cycle, and DNA

Damage

Apoptosis and cell prolieration assays are particularly useul

or basic cancer research and drug discovery. Comparing

data across dierent experiments can be challenging due

to variability introduced by sample handling, timing, and

variability within the sample.

Multicolor low cytometry addresses these challenges andis an excellent tool to study apoptosis and cell prolieration.

Relevant markers can be combined with cell phenotyping

markers to look at events within subpopulations o cells.

Antibodies to phosphoproteins can be used to examine

phosphorylation events.

CleavedParp-AlexaFluo

r6

47

BrdU - FITC

102 103 104 105

102

103

104

105

Untreated

3% 1%

64% 32%

102 103 104 105

102

103

104

105

20 M Camptothecin

26% 10%

50% 14%

102 103 104 105

102

103

104

105

10 M Camptothecin

27% 9%

50% 14%

102 103 104 105

102

103

104

105

5 M Camptothecin

15% 6%

60% 19%

102 103 104 105

102

103

104

105

2.5 M Camptothecin

7% 3%

67% 23%

A

B

A

B

Immunofluorescence of cleaved PARP

HeLa cells grown were either let untreated (A) or treated with staurosporine (1.0

mM, 4 h) to induce apoptosis (B). Cells were then ixed with 3.7% ormaldehyde(15 min on ice), then permeabilized in 0.25% Triton X-100/3% BSA/PBS (15 min

on ice). Cells were then washed twice with 3% BSA/PBS and stained with 4 L/mL o

FITC-labeled anti-PARP in 3% BSA/PBS (1 h at RT). Cells were washed twice with 3%

BSA/PBS and then visualized by immunoluorescence microscopy. A and B represent

phase correlates o A and B, respectively. The results indicate that untreated cells

were primarily negative or cleaved PARP (A), whereas a signiicant percentage o

the staurosporine-treated population is positive or cleaved PARP.



ServicesBD Biosciences instruments and reagents are backed

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cytometry instrumentation with trusted, certiied reagents,

and advanced applications. The BD Biosciences tools enable

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Researchers come to BD Biosciences not only or quality

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For example, our website, bdbiosciences.com, now

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References

1. Lodish H, D. Baltimore A, Berk S L, Zipursky P,Matsudaira J, Darnell, eds. Cell organization,subcellular structure, and cell division. 1995.Molecular Cell Biology. Third Edition. W. H.Freeman and Company, New York, pp. 141-188.

2. Prez-Cadaha B, Drobic B, Davie JR. H3phosphorylation: dual role in mitosis andinterphase. Biochem Cell Biol. 2009;87:695-709.

3. Hedrick SM, Chen IL, Alves BN. Intertwinedpathways o programmed cell death in immunity.Immunol Rev. 2010;236:41-53. Review.

4. Salvesen GS, Riedl SJ. Caspase mechanisms. Adv ExpMed Biol. 2008;615:13-23. Review.

5. Thornberry NA, Chapman KT, Nicholson DW.Determination o caspase specifcities using apeptide combinatorial library. Methods Enzymol.2000;322:100-110.

6. Buggins AG, Pepper CJ. The role o Bcl-2 amilyproteins in chronic lymphocytic leukaemia. LeukRes. 2010;34:837-842. Epub 2010 Mar 31. Review.

7. Russo M, Mupo A, Spagnuolo C, Russo GL.Exploring death receptor pathways as selectivetargets in cancer therapy. Biochem Pharmacol.2010;80(5):674-682. Epub 2010 Mar 17. Review.

8. Boulares AH, Yakovlev AG, Ivanova V, et al. Roleo poly(ADP-ribose) polymerase (PARP) cleavagein apoptosis. Caspase 3-resistant PARP mutantincreases rates o apoptosis in transected cells.J Biol Chem. 1999;274:22932-22940.

9. Rouleau M, Patel A, Hendzel MJ, Kaumann SH,Poirier GG. PARP inhibition: PARP1 and beyond.Nat Rev Cancer. 2010;10:293-301. Epub 2010 Mar Review.

10. Tanaka T, Huang X, Halicka HD, et al. Cytometry oATM activation and histone H2AX phosphorylatioto estimate extent o DNA damage induced byexogenous agents. Cytometry A. 2007;71:648-661Review.


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