Human/Mouse/Rat HIF-1 alpha/HIF1A Antibody

Detection of Human HIF‑1 alpha/HIF1A by Simple Western^TM.

Simple Western lane view shows lysates of A549 human lung carcinoma cell line untreated (-) or treated (+) with Hypoxia (1% O₂), loaded at 0.2 mg/mL. A specific band was detected for HIF-1a/HIF1A at approximately 115 kDa (as indicated) using 5 µg/mL of Goat Anti-Human/Mouse/Rat HIF-1a/HIF1A Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1935) followed by 1:50 dilution of HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF109). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.

Detection of Mouse HIF-1 alpha by Simple Western

Ten days of IH increases hippocampal HIF1a and disrupts Barnes maze performance in wild-type mice but not in HIF1a+/−. A, left, Representative digitized Western blotting images for HIF1a (103 kDa) and PCNA (40 kDa) in hippocampal nuclear protein fractions from control (n = 4) and IH10 (n = 4). Right, Quantification of HIF1a protein normalized to PCNA revealed that nuclear HIF1a was increased in IH10 when compared with control (p = 0.019). B, Total latency to exit the Barnes maze during three training sessions in control (n = 10) and in IH10 (n = 11). Each blue (control) and red (IH10) line represents an individual performance during training. Training to the exit was conducted over three sessions. Each session was separated by 24 hours. C, Left, During the probe trial, the distance traveled to initially enter the exit zone was shorter in control when compared with IH10 (p = 0.048). Right, Latency to initial entry was smaller in control as well (p = 0.034). D, Heat maps of the mean entry probability across all false exits (1–19) and the exit zone during probe trial for the control and IH10. Comparison of entry probability into the exit zone during the probe trial reveals that control has a greater probability for entering the exit zone when compared with IH10 (p = 0.004). E, Left, Representative digitized Western blotting images HIF1a and PCNA in hippocampal nuclear protein fractions from 0-HIF1a+/− (n = 4) and 10-HIF1a+/− (n = 4). Right, Quantification of HIF1a protein normalized to PCNA revealed that nuclear HIF1a is similar between 0-HIF1a+/− and 10-HIF1a+/− (p = 0.84). F, Total latency to exit the Barnes maze during three training sessions in 0-HIF1a+/− (n = 7) and in 10-HIF1a+/− (n = 8). Each gray (0-HIF1a+/−) and yellow (10-HIF1a+/−) line represents an individual performance during training. All experimental groups exhibit decreased total latency over the course of training. G, Left, In HIF1a+/−, the distance initial to initial entry into the exit zone was similar between 0-HIF1a+/− and 10-HIF1a+/− (p = 0.55). Right, Latency to initial entry into the exit zone during the probe trial were similar between 0-HIF1a+/− and 10-HIF1a+/− (p = 0.39). H, Heat maps of the mean entry probability into all zones during the probe trial for 0-HIF1a+/− and 10-HIF1a+/−. Entry probability was similar between 0-HIF1a+/− and 10-HIF1a+/− (p = 0.21); *p < 0.05; **p < 0.01; N.S., p > 0.05. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32493757), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human HIF-1 alpha by Western Blot

The DU145FP cells express higher levels of cancer stemness related genes. (A) Transcriptome analyses of prostate cancer stemness-related markers. The Log2 transformed ratio of FPKM values (e.g., FP/WT) are indicated by color-coded index bars. (B) Cells were stained with fluorophore conjugated monoclonal antibodies against CD44 and EPCAM, the antibody against integrin alpha2 beta3 without fluorophore conjugation was detected by using secondary antibody conjugated with fluorophore, and stained cells were analyzed by flow cytometry. The scatter plots and histograms show representative results. CTRL denote cell stained with isotype control antibody or only secondary antibody; ST denote cell stained with antibody against EPCAM and integrin alpha2 beta3; US denote unstained cells. (C) Immunoblotting confirmation of the protein expression of A-tubulin, GAPDH, SOX9, SLUG, HIF1A, HIF2A, CD44 and integrin alpha2 beta3. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/28698547), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse HIF-1 alpha by Simple Western

Ten days of IH increases hippocampal HIF1a and disrupts Barnes maze performance in wild-type mice but not in HIF1a+/−. A, left, Representative digitized Western blotting images for HIF1a (103 kDa) and PCNA (40 kDa) in hippocampal nuclear protein fractions from control (n = 4) and IH10 (n = 4). Right, Quantification of HIF1a protein normalized to PCNA revealed that nuclear HIF1a was increased in IH10 when compared with control (p = 0.019). B, Total latency to exit the Barnes maze during three training sessions in control (n = 10) and in IH10 (n = 11). Each blue (control) and red (IH10) line represents an individual performance during training. Training to the exit was conducted over three sessions. Each session was separated by 24 hours. C, Left, During the probe trial, the distance traveled to initially enter the exit zone was shorter in control when compared with IH10 (p = 0.048). Right, Latency to initial entry was smaller in control as well (p = 0.034). D, Heat maps of the mean entry probability across all false exits (1–19) and the exit zone during probe trial for the control and IH10. Comparison of entry probability into the exit zone during the probe trial reveals that control has a greater probability for entering the exit zone when compared with IH10 (p = 0.004). E, Left, Representative digitized Western blotting images HIF1a and PCNA in hippocampal nuclear protein fractions from 0-HIF1a+/− (n = 4) and 10-HIF1a+/− (n = 4). Right, Quantification of HIF1a protein normalized to PCNA revealed that nuclear HIF1a is similar between 0-HIF1a+/− and 10-HIF1a+/− (p = 0.84). F, Total latency to exit the Barnes maze during three training sessions in 0-HIF1a+/− (n = 7) and in 10-HIF1a+/− (n = 8). Each gray (0-HIF1a+/−) and yellow (10-HIF1a+/−) line represents an individual performance during training. All experimental groups exhibit decreased total latency over the course of training. G, Left, In HIF1a+/−, the distance initial to initial entry into the exit zone was similar between 0-HIF1a+/− and 10-HIF1a+/− (p = 0.55). Right, Latency to initial entry into the exit zone during the probe trial were similar between 0-HIF1a+/− and 10-HIF1a+/− (p = 0.39). H, Heat maps of the mean entry probability into all zones during the probe trial for 0-HIF1a+/− and 10-HIF1a+/−. Entry probability was similar between 0-HIF1a+/− and 10-HIF1a+/− (p = 0.21); *p < 0.05; **p < 0.01; N.S., p > 0.05. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32493757), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human HIF-1 alpha/HIF1A by Immunocytochemistry/ Immunofluorescence

Staining of HIF1A in HIF1A KO and WT HUVECs. (a–d) One representative image of the stained HIF1A KO and WT HUVECs, which were stained with Hoechst (blue), anti-actin (green), and anti-HIF1A (red). (a) WT HUVECs cultured in normoxic conditions. (b) WT HUVECs cultured in hypoxic conditions. (c) HIF1A KO HUVECs cultured in normoxic conditions. (d) HIF1A KO HUVECs cultured in hypoxic conditions. (e) Quantification of the HIF1A signal intensity (MFI) in the nucleus and the cytoplasm in WT and KO HUVECs in normoxic and hypoxic conditions, which is compared by two-way ANOVA multiple comparisons test, *** p ≤ 0.001. The experiment was performed with one biological replicate at p. 3 (n = 3) and five images were taken per well. Scale bar 10 µm. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36671408), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human HIF-1 alpha/HIF1A by Immunocytochemistry/ Immunofluorescence

Staining of HIF1A in HIF1A KO and WT HUVECs. (a–d) One representative image of the stained HIF1A KO and WT HUVECs, which were stained with Hoechst (blue), anti-actin (green), and anti-HIF1A (red). (a) WT HUVECs cultured in normoxic conditions. (b) WT HUVECs cultured in hypoxic conditions. (c) HIF1A KO HUVECs cultured in normoxic conditions. (d) HIF1A KO HUVECs cultured in hypoxic conditions. (e) Quantification of the HIF1A signal intensity (MFI) in the nucleus and the cytoplasm in WT and KO HUVECs in normoxic and hypoxic conditions, which is compared by two-way ANOVA multiple comparisons test, *** p ≤ 0.001. The experiment was performed with one biological replicate at p. 3 (n = 3) and five images were taken per well. Scale bar 10 µm. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36671408), licensed under a CC-BY license. Not internally tested by R&D Systems.