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REACTIVITY MW (kDa)

Figure 1. Target map of the PathScan® Stress and Apoptosis Signaling Antibody Array Kit (Fluorescent Readout) #12923.

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Figure 2. HeLa cells were grown to 90% confluency and then either untreated (A, left panel) or treated with Staurosporine #9953 (1 μM, 3.5 hr; A, right panel). Cell extracts were prepared and analyzed using the PathScan® Stress and Apoptosis Signaling Antibody Array Kit (Fluorescent Readout) #12923. Panel A shows images that were acquired using the LI-COR® Biosciences Odyssey® imaging system. Panel B shows quantified fluorescence intensity (RFU) for each target in the presence or absence of staurosporine. Relative fluorescent intensities were quantified as pixel intensities using the LI-COR® Image Studio v2.0 array analysis software.

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Figure 3. HT-29 cells were grown to 80% confluency and then either untreated (A, left panel) or UV-irradiated and allowed to recover for 60 minutes (A, right panel). Cell extracts were prepared and analyzed using the PathScan® Stress and Apoptosis Signaling Antibody Array Kit (Fluorescent Readout) #12923. Panel B shows quantified fluorescence intensity (RFU) for each target in the presence or absence of UV-irradiation. Relative fluorescent intensities were quantified as pixel intensities using the LI-COR® Image Studio v2.0 array analysis software.

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Figure 4. HeLa cells were grown to 90% confluency and then either untreated (A, left panel) or treated with Human Tumor Necrosis Factor-α (hTNF-α) #8902 (100 ng/ml, 20 min; A, right panel). Cell extracts were prepared and analyzed using the PathScan® Stress and Apoptosis Signaling Antibody Array Kit (Fluorescent Readout) #12923. Panel A shows images that were acquired using the LI-COR® Biosciences Odyssey® imaging system. Panel B shows quantified fluorescence intensity (RFU) for each target in the presence or absence of hTNF-α. Relative fluorescent intensities were quantified as pixel intensities using the LI-COR® Image Studio v2.0 array analysis software.

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Figure 5. HeLa cells were grown to 90% confluency and serum starved overnight. Cells were then either untreated (A, left panel) or treated with Human Transforming Growth Factor β3 (hTGF-β3) #8425 (100 ng/ml, 20 min; A, right panel). Cell extracts were prepared and analyzed using the PathScan® Stress and Apoptosis Signaling Antibody Array Kit (Fluorescent Readout) #12923. Panel A shows images that were acquired using the LI-COR® Biosciences Odyssey® imaging system. Panel B shows quantified fluorescence intensity (RFU) for each target in the presence or absence of hTGF-β3. Relative fluorescent intensities were quantified as pixel intensities using the LI-COR® Image Studio v2.0 array analysis software.

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Figure 6. Various cell lysates were analyzed using the PathScan® Stress and Apoptosis Signaling Antibody Array Kit (Fluorescent Readout) #12923. Images were acquired using the LI-COR® Biosciences Odyssey® imaging system. Fluorescence intensity was quantified using the LI-COR® Image Studio v2.0 array analysis software. Heatmap analysis was generated using MultiExperiment Viewer (MeV) analysis software using the raw fluorescence intensity values.

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Image
Product Includes Quantity
Array Slides - Stress and Apoptosis Array 2 Ea
16-Well Gasket 2 Ea
Sealing Tape 2 sheets
20X Array Wash Buffer 15 ml
Array Blocking Buffer 5 ml
Array Diluent Buffer 15 ml
Detection Ab Cocktail 10X - Stress and Apoptosis Array 300 µl
DyLight 680TM-linked Streptavidin (10X) 300 µl
PathScan® Sandwich ELISA Lysis Buffer (1X) 7018 30 ml

Product Usage Information

Storage: Kit should be stored at 4°C with the exception of Lysis Buffer, which is stored at –20°C (packaged separately).

Product Description

The PathScan® Stress and Apoptosis Signaling Antibody Array Kit (Fluorescent Readout) uses glass slides as the planar surface and is based upon the sandwich immunoassay principle. The array kit allows for the simultaneous detection of 19 signaling molecules that are involved in the regulation of the stress response and apoptosis. Target-specific capture antibodies have been spotted in duplicate onto nitrocellulose-coated glass slides. Each kit contains two slides allowing for the interrogation of 32 different samples and the generation of 608 data points in a single experiment. Cell lysate is incubated on the slide followed by a biotinylated detection antibody cocktail. Streptavidin-conjugated DyLight® 680 is then used to visualize the bound detection antibody. A fluorescent image of the slide can then be captured with a digital imaging system and spot intensities quantified using array analysis software.


Specificity / Sensitivity

PathScan® Stress and Apoptosis Signaling Antibody Array Kit (Fluorescent Readout) detects the target proteins as specified on the Array Target Map. No substantial cross-reactivity has been observed between targets. This kit is optimized for cell lysates diluted to a total protein concentration between 0.2 and 1 mg/ml (see kit protocol). All capture antibodies have been validated for human and mouse-derived samples.


Cell death can occur due to a variety of circumstances including nutrient deprivation, inability to generate or store the energy required for metabolic functions, or deleterious environment that causes irreparable damage. Cells integrate multiple signals from a variety of sources before following either pro- or anti-apoptotic pathways. These signals can often carry conflicting information. Assessing the net effect of these processes in cell populations can be achieved by monitoring changes in a number of key signaling components. The caspase-3 and caspase-7 proteases exert a pro-apoptotic function through cleavage of multiple cellular targets. Caspase-3 and caspase-7 are activated by cleavage at Asp175 and Asp198, respectively. PARP is a DNA repair and apoptosis enzyme that is inactivated by cleavage at Asp214 by caspase-3 or caspase-7. HSP27 is a mediator of cell stress that confers resistance to adverse environmental conditions. HSP27 is activated by phosphorylation at Ser82. Chk1 and Chk2 kinases act downstream of ATM/ATR and play an important role in DNA damage checkpoint control. Activation of Chk1 and Chk2 involve phosphorylation at Ser345 and Thr68, respectively. Tumor suppressor p53 plays an important role in cellular response to DNA damage. p53 is phosphorylated at Ser15 by ATM/ATR or DNA-PK leading to its accumulation. Smad2 is a key mediator of TGF-β signaling. Stimulation by TGF-β leads to Smad2 phosphorylation at Ser465/467 and translocation of Smad2 into the nucleus. The outcome of TGF-β signaling is context dependent and can either induce apoptosis or contribute to tumor cell metastasis. Activation of NF-κB/Rel occurs through a proteasome-mediated degradation of IκBα. The inhibitor IκBα is targeted to the proteasome via phosphorylation of IκBα at Ser32 and Ser36. NF-κB activation is triggered by a diverse group of extracellular signals promoted by inflammatory cytokines, growth factors, and chemokines. TAK1 is a kinase that can be activated by TGF-β, bone morphogenetic proteins and other cytokines. Activated TAK1 phosphorylates MKK4, MKK3/6, and NIK. Phosphorylation of TAK1 at Ser412 is one of the mechanisms that regulate the levels of its activation. Cellular stress such as viral infection, endoplasmic reticulum stress, and amino acid deprivation leads to phosphorylation of eIF2α. Phosphorylation of eIF2α at Ser51 in response to cellular stress leads to a reduction of protein synthesis. The ERK1 and ERK2 MAP kinases are major signaling nodes that have many substrates and primarily transmit growth and proliferation signals. The ERK MAP kinase is activated by a dual phosphorylation of Thr202 and Tyr204. p38 MAPK and SAPK/JNK MAP kinases are activated through a similar dual phosphorylation mechanism in response to pro-inflammatory cytokines and genotoxic stress. Akt is activated by stimulation of growth-factor receptors and primarily promotes anabolic growth and survival signals via targeting its broad array of substrates. Akt phosphorylates Bad at Ser136 and inhibits its ability to induce apoptosis. Survivin is an anti-apoptotic protein that is highly expressed during fetal development and cancer cell malignancy. Survivin binds and inhibits caspase-3, controlling the cell cycle by inhibiting apoptosis and promoting cell division. α-tubulin is a building block of microtubules that are present in all eukaryotic cells. The levels of the globular α-tubulin are considered to remain relatively constant. Therefore, assessing the relative levels of α-tubulin may assist with signal normalization between the various samples.


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Entrez-Gene Id 207, 208, 10000, 572, 836, 840, 1111, 11200, 1965, 5595, 5594, 3315, 4792, 142, 1432, 5600, 5603, 6300, 7157, 5599, 4087, 332, 6885, 10376
Swiss-Prot Acc. P31749, P31751, Q9Y243, Q92934, P42574, P55210, O14757, O96017, P05198, P27361, P28482, P04792, P25963, P09874, Q16539, Q15759, O15264, P53778, P04637, P45983, Q15796, O15392, O43318, P68363

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