Human Hif1a + BNIP3 Hypoxia In Cell ELISA Kit (IR)

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Human Hif1a + BNIP3 Hypoxia In Cell ELISA Kit (IR) (AB129733)

Antibody specificity demonstrated by immunocytochemistry. Primary antibodies used in this assay kit were validated by staining HeLa cells +/- treatment with 1mM DFO (24h) and imaged by fluorescent microscopy. HIF1 alpha staining is absent in untreated cells and induced by DFO treatment. HIF1 alpha localizes to the nucleus (as seen by co-localization with the DNA stain DAPI) as expected.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Human Hif1a + BNIP3 Hypoxia In Cell ELISA Kit (IR) (AB129733)

Antibody specificity demonstrated by immunocytochemistry. Primary antibodies used in this assay kit were validated by staining HeLa cells +/- treatment with 1mM DFO (24h) and imaged by fluorescent microscopy. BNIP3 staining is nearly undetectable in untreated HeLa cells but is induced by DFO treatment. BNIP3 appears to has a mitochondrial staining pattern in the DFO-treated samples.

• In-Cell ELISA

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In-Cell ELISA - Human Hif1a + BNIP3 Hypoxia In Cell ELISA Kit (IR) (AB129733)

Sample experiment using ab129733 on HeLa cells treated with a titration of DFO. HeLa cells were seeded to an amine coated 96-well microplate and the following day treated with a titration of DFO. After 24h of DFO exposure, the cells were fixed and stained as described in the protocol and the normalized data is presented here +/- SD (as described in the protocol and data analysis sections). BNIP3 levels are increased with DFO concentrations ≥10µM.

• In-Cell ELISA

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In-Cell ELISA - Human Hif1a + BNIP3 Hypoxia In Cell ELISA Kit (IR) (AB129733)

Coefficient of variation for the experiment described in Figure 1.

• In-Cell ELISA

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In-Cell ELISA - Human Hif1a + BNIP3 Hypoxia In Cell ELISA Kit (IR) (AB129733)

Sample experiment using ab129733 on HeLa cells treated with a titration of DFO. HeLa cells were seeded to an amine coated 96-well microplate and the following day treated with a titration of DFO. After 24h of DFO exposure, the cells were fixed and stained as described in the protocol and the normalized data is presented here +/- SD (as described in the protocol and data analysis sections). HIF1 alpha results show DFO concentrations ≥10µM induce HIF1A protein levels in a dose dependent manner.

• WB

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Western blot - Human Hif1a + BNIP3 Hypoxia In Cell ELISA Kit (IR) (AB129733)

Antibody specificity demonstrated by Western Blot. Primary antibodies used in this assay kit were validated by Western Blot using HeLa cell lysates that had been treated with a dose titration of DFO as indicated. (A) The HIF1 alpha band (indicated by arrow) is absent in untreated cells and induced by DFO. (B) Similarly, BNIP3 levels are increased by DFO treatment in a dose-dependent manner.

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