Apoptosis Assays for Advanced High-Throughput Flow Cytometry

Apoptosis is an essential process for normal tissue development and homeostasis by which cells undergo timely programmed cell death. Aberrations in apoptotic signaling are implicated in a range of human pathologies including cancer, autoimmune disease and neurodegeneration. Induction of apoptosis leads, in most cases, to the depolarization of mitochondria, the activation of caspases and plasma membrane alterations. The ability to simultaneously measure a multitude of apoptotic markers allows confirmation of pathway and insight into mechanism of action over time.


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Assay Concept

Introducing Apoptosis Assays for Flow Cytometry

Figure 1. Apoptotic markers measured by the iQue® Human 4-Plex Apoptosis Kit. 1) Mitochondrial depolarization, 2) Caspase 3/7, 3) Annexin V and 4) Non-viable. The kit components can be utilized either separately or combination in a no wash format. Apoptosis reagents can also be multiplexed with other iQue® reagents in no wash assays.

Key Advantages

Key Advantages of Apoptosis Assays

  • Detect and confirm apoptosis through different pathways using the mix and match reagents
  • Gain a robust determination of apoptosis progression and insight into mechanism of action
  • Simple mix-and-read 96 and 384-well protocol with no washing and no fixing

Detect and Confirm Apoptosis Through Different Pathways Using the Mix and Match Reagents

Figure 2. Cytotoxic and cytostatic compounds produce different cell health profiles. Jurkats (1e6/mL) treated with a range of concentrations of 4 compounds for 24 hours. Before analysis with iQue®️ Human 4-Plex Apoptosis Kit.

(A) Caspase 3/7 expressing cells are shown in a heatmap from iQue Forecyt®️.

(B,C,D,E) Depolarized mitochondria, Caspase 3/7, Annexin V and Non-viable (respectively) cell percentages are shown.

Gain a Robust Determination of Apoptosis Progression and Insight Into Mechanism of Action

Figure 3. Cell health markers show a time dependent increase when treated with an apoptosis inducing drug.
Jurkats (1e6/mL) treated with a range of concentrations of Staurosporin. 20 µL samples were taken and analyzed after 2, 6 and 24 hours of treatment with 3 apoptosis reagents. (A) Heat map of Caspase positive cell (%) from 2 hours, (B) Mitochondrial depolarization, (C) Caspase and (D) Annexin V

Simple Mix-and-Read 96 and 384-Well Protocol With No Washing and No Fixing

Figure 4. Schematic of the simple, mix and read apoptosis assay.

Ordering Information

Ordering Information

iQue®️ Human 4-Plex Apoptosis Kit

Platform: Compatible with iQue®️ 3 BR/VBR & iQue®️ Plus BR/VBR

Available Sizes

Catalog Numbers

1 x 384

90053

5 x 384

90054

20 x 384

90155

50 x 384

90156


Individual Apoptosis Reagents

Platform: Compatible with iQue®️ 3 BR/VBR & iQue®️ Plus BR/VBR
ProductDetection Channel

Available Size

Catalog Number

iQue® Human Caspase 3/7

B/Green

5 x 384
20 x 384
50 x 384

91034
91035
91036

iQue® Human Annexin V

B/Yellow

5 x 384
20 x 384
50 x 384

91030
91031
91032

iQue® Mitochondrial Membrane Potential

R/Red

5 x 384
20 x 384
50 x 384

91038
91039
91040

iQue® Cell Membrane Integrity

B/Red

5 x 384
20 x 384
50 x 384

90346
90347
90348


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Resources

Literature and Documentation

Product Guide

iQue®️ Human 4-Plex Apoptosis Kit

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Poster: ELRIG 2014

High throughput multiplexed apoptosis assays using the Labcyte Echo Liquid Handler and the Intellicyt iQue Screener HD

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Poster: SLAS 2014

Rapid profiling of compounds for cell cycle and apoptosis: using single micro-scale compound treatments for multiple multiplexed assays

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Infographic

An advanced flow cytometry approach to assessing cell health and viability

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Application Note

Development of a no-wash 4-plex apoptosis screening kit and validation for phenotypic screening

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Application Note

Evaluating cellular progression through apoptosis via multiple different pathways

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