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NC-3000 Annexin V Assay– For easy estimation of apoptosis

Translocalisation of phosphatidylserine (PS) from the inner to the outer membrane layer happens early in apoptosis andprecedes other apoptotic events such as loss of membrane in-tegrity and DNA fragmentation. Annexins are group of cellular proteins that bind to phospholipids in a calcium-dependentmanner. A member of this group, Annexin V, has proven to be auseful tool in detecting apoptotic cells since it has a high afn-ity for PS. Using uorescently labeled Annexin V the externaliza-tion of PS can easily be detected. Annexin V may also bind toPS on late apoptotic and necrotic cells but as the membraneintegrity on these cells has been lost, these cells can be distin-guished from early apoptotic cells by the use of the impermeantdye propidium iodide.

Key Beneftsof the NC-3000 Annexin V Assay

STEP 1: Add Annexin V FITC conjugate

and Hoechst 33342Incubate 15 minutes

STEP 2: Wash and resuspend cellsin PI-containing buffer

Image cytometry

Introducing the NucleoCounter®

NC-3000™

 - Performing Automated Image Cytometry 

 Viability and Cell Count |  Two-step Cell Cycle |  Cell Cycle of Fixed Cells |  Apoptosis Detection  Mitochondrial Potential | Cell Vitality |  DNA Fragmentation |  GFP Transfection Efciency |  Annexin V | and more 

 Information about both earlyapoptotic and late apoptotic/necroticcells provided

 Fast automated single cell analysis

  Acquisition, analysis and datapresentation in one simple step

 Well-recognised assay

 User friendly protocol withpredened settings

 Standardized results – even withdifferent users

 No calibration required

 A  C Q U I S I T I O N ,

 A  N A  L Y S I S  A  N D

 D A  T A   P R E S E N T A  T

 I O N

 I N  O N E

 S I M P L E  S T E P

8/4/2019 Application Flyer Annexin v Assay HP

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Principle

ChemoMetec A/S

Gydevang 43

DK-3450 Allerød

Denmark

Phone (+45) 48 13 10 20

Fax (+45) 48 13 10 21

Mail contact@chemometec.com

Web www.chemometec.com

Using uorescence microscopy and image analysis the NC-3000 system auto-mates detection of apoptotic cells based on phosphatidylserine externaliza-tion. Cells are stained with Hoechst 33342, PI and FITC labeled Annexin V.

Hoechst 33342 stains the total cell population, while Annexin V stains apop-totic and necrotic cells. Early apoptotic cells exclude PI, while late stageapoptotic and necrotic cells stain positively for both Annexin V and PI.

For more information, please visit  www.chemometec.com/assays

 Viability and Cell Count | Two-step Cell Cycle | Cell Cycle of Fixed Cells |  Apoptosis Detection | Cell VitalityMitochondrial Potential | DNA Fragmentation | GFP Transfection Efciency | Annexin V | and more 

NC-3000 Annexin V Assay- For easy estimation of apoptosis

 Jurkat cells stained with Hoechst-33342, Annexin V FITC conjugate and propidium iodide (PI). Cells were grown in the absence(upper left) or in the presence of 10 µM camptothecin (CPT) for 5 hours at 37°C (upper right). Cells were stained and analysed onNC-3000 using the Annexin V protocol. In this example 2% of the untreated Jurkat cells are early apopotic, whereas 40% of theCPT treated Jurkat cells are in early apoptosis. Lower panel shows the images acquired for the two samples.

Annexin V FITC Intensity Annexin V FITC Intensity

   P   I   I   n   t   e   n   s    i   t   y

   P   I   I   n   t   e   n   s    i   t   y

Untreated Jurkat cells CPT treated Jurkat cells