Antibody data
- Antibody Data
- Antigen structure
- References [1]
- Comments [0]
- Validations
- Western blot [2]
- Immunocytochemistry [1]
- Immunohistochemistry [1]
- Flow cytometry [1]
- Other assay [2]
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Validation data
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- Product number
- 38-5600 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- Anti-IRAK1 Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Reactivity
- Human
- Host
- Rabbit
- Isotype
- IgG
- Vial size
- 100 µg
- Concentration
- 0.25 mg/mL
- Storage
- -20°C
Submitted references Myeloid Differentiation Primary Response 88-Cyclin D1 Signaling in Breast Cancer Cells Regulates Toll-Like Receptor 3-Mediated Cell Proliferation.
Singh A, Devkar R, Basu A
Frontiers in oncology 2020;10:1780
Frontiers in oncology 2020;10:1780
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of (A) MCF-7 and (B) MDA-MB231 cell lysates using Rb anti IRAK-1 (Product # 38-5600)
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of IRAK1 was performed by loading 20 µg of MDA-MB-231 (lane1) and K562 (lane2) cell lysate using Novex® NuPAGE® 12 % Bis-Tris gel (Product # NP0342BOX), XCell SureLock™ Electrophoresis System (Product # EI0002), Novex® Sharp Pre-Stained Protein Standard (LC5800), and iBlot® 2 Dry Blotting System (IB21001). Proteins were transferred to a nitrocellulose membrane and blocked with 5 % skim milk for 1 hour at room temperature. IRAK1 was detected at ~ 75 kDa using IRAK1 Rabbit Polyclonal Antibody (Product # 38-5600) at 1-2 µg/mL in 5 % skim milk at 4°C overnight on a rocking platform. Goat Anti-Rabbit IgG - HRP Secondary Antibody (G21234) at 1:5000 dilution was used and chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescent analysis of IRAK-1 Antibody was done on 70% confluent log phase HeLa cells. The cells were fixed with 4% paraformaldehyde for 15 minutes, permeabilized with 0.25% Triton™ X-100 for 10 minutes, and blocked with 5% BSA for 1 hour at room temperature. The cells were labeled with IRAK-1 Antibody (Product # 38-5600) at 1µg/mL in 1% BSA and incubated for 3 hours at room temperature and then labeled with Alexa Fluor 488 Goat Anti-Rabbit IgG Secondary Antibody (Product # A-11008) at a dilution of 1:400 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Alexa Fluor 594 Phalloidin (Product # A12381). Panel d is a merged image showing Nuclear localization. Panel e is a no primary antibody control. The images were captured at 40X magnification.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemistry analysis of IRAK1 showing staining in the cytoplasm of paraffin-embedded human kidney tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a IRAK1 polyclonal antibody (Product # 38-5600) diluted in 3% BSA-PBS at a dilution of 1:100 overnight at 4ºC in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Flow cytometry analysis of IRAK1 was done on HeLa cells. Cells were fixed with 70% ethanol for 10 minutes, permeabilized with 0.25% Triton™ X-100 for 20 minutes, and blocked with 5% BSA for 30 minutes at room temperature. Cells were labeled with IRAK1 Rabbit Polyclonal Antibody (385600, red histogram) or with rabbit isotype control (pink histogram) at 3-5 ug/million cells in 2.5% BSA. After incubation at room temperature for 2 hours, the cells were labeled with Alexa Fluor® 488 Goat Anti-Rabbit Secondary Antibody (A11008) at a dilution of 1:400 for 30 minutes at room temperature. The representative 10,000 cells were acquired and analyzed for each sample using an Attune® Acoustic Focusing Cytometer. The purple histogram represents unstained control cells and the green histogram represents no-primary-antibody control.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Figure 5 Western blotting for the expression of signaling protein. (A,B) Cell lysate were collected and subjected to Western blot assay to estimate the level of expression of interleukin 1 receptor-associated kinase 1 (IRAK1), transforming growth factor beta-activated kinase 1 (TAK1), TGF-beta-activated kinase 1 (TAB1), TNF receptor-associated factor 6 (TRAF6), and cyclin D1. (C,D) Expression of pIRAK1 and pTAK1. beta-actin was used as loading control. The respective bar graphs are presented as densitometry analysis as mean +- SD of experiments ( p < 0.05 is treated as significant).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Figure 6 Immunoprecipitation showing the involvement of the signaling complex. (A) Signaling complex of IRAK1/TRAF6 was immunoprecipitated with antibodies against IRAK1 followed by Western blotting with anti-TRAF6 and anti-IRAK1 antibodies. (B) Signaling complex of pIRAK1/TAK1 was immunoprecipitated with antibodies against pIRAK1 followed by Western blotting with anti-TAK1 and anti-pIRAK1 antibodies. (C) Signaling complex TAB1-TRAF6-TAK1 was immunoprecipitated with antibodies against pTAK1 followed by Western blotting using anti-TRAF6, TAB1, and pTAK1 antibodies. (D) Signaling complex TAB1-TRAF6-TAK1 was immunoprecipitated with antibodies against TRAF6 followed by Western blotting analysis using anti-TAK1, TAB1, and TRAF6 antibodies.