Western blot was performed on blood samples. Blood samples were incubated with primary rabbit polyclonal antibodies and mouse monoclonal antibodies, anti-cyclooxygenase 1 (COX-1) (ab109025) (Abcam, Cambridge, MA, USA), anti-TGF beta 1 (ab2486) (Abcam, Cambridge, MA, USA), anti-Smad3 (ab40854) (Abcam, Cambridge, MA, USA), and anti-cyclooxygenase 2 (COX-2) (Cat. No. 12282S) (Cell Signaling Technology, Danvers, MA, USA). Secondary antibodies included goat anti-rabbit IgG (1: 5000) (Multisciences, Hangzhou, China), and goat anti-mouse IgG (1: 5000) (Multiscience, Hangzhou, China). Immunostains were visualized using electrochemiluminescence (ECL) and Western blot (Beyotime Biotechnology Co. Ltd., Shanghai), and quantified using the ChemiDoc XRS+ imaging system (Bio-Rad Labs, Hercules, CA, USA).
Ab2486
Manufactured by Abcam
Sourced in United States
Ab2486 is a primary antibody produced in rabbit and targeted against a specific antigen. It is designed for use in various laboratory techniques, including immunohistochemistry and western blotting.
Automatically generated - may contain errors
Lab products found in correlation
Phenyl methyl sulfonyl fluoride (pmsf), by Beyotime (3 mentions)
Ab142349, by Abcam (3 mentions)
Ecl reagent, by Merck Group (3 mentions)
Ab6672, by Abcam (2 mentions)
Ab133462, by Abcam (2 mentions)
Ab40854, by Abcam (1 mentions)
Anti cyclooxygenase 2 cox 2, by Abcam (1 mentions)
Chemidoc xrs imaging system, by Bio-Rad (1 mentions)
Ab109025, by Abcam (1 mentions)
Goat anti mouse igg, by MultiSciences Biotech (1 mentions)
Goat anti rabbit igg, by MultiSciences Biotech (1 mentions)
Ab137133, by Abcam (1 mentions)
Ab652, by Abcam (1 mentions)
Ab92498, by Abcam (1 mentions)
Ab2571, by Abcam (1 mentions)
Ab32359, by Abcam (1 mentions)
Bms213 2, by Thermo Fisher Scientific (1 mentions)
Hrp conjugated anti rabbit or anti mouse antibodies, by Santa Cruz Biotechnology (1 mentions)
Ab7217, by Abcam (1 mentions)
Ab51177, by Abcam (1 mentions)
4 protocols using ab2486
1
Western Blot Analysis of Inflammatory Markers
2
Western Blot Analysis of HIF1α and mTOR
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For western blotting, proteins were extracted from cultured cells using RIPA buffer containing PMSF (Beyotime). Equal amounts of protein (100 μg) were separated by 7.5%/12.5% SDS‐PAGE and transferred to a PVDF membrane. Primary polyclonal Abs targeting HIF1a (ab92498), REDD1 (ab63059), mTOR Ser2481 (ab137133), mTOR Ser2448 (ab2486), mTOR (ab142349), and GLUT1 (ab652) were purchased from Abcam. The secondary Abs used were anti‐rabbit HRP‐linked (Abcam). The blots were developed using ECL reagent (Millipore). An equal amount of protein loading in each lane was confirmed using the β‐actin Ab. ImageJ software (National Health Institute US) was used to quantify the integrated density of the band.
3
Western Blot Analysis of Signaling Pathways
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For western blotting, proteins were extracted from tissues or cultured cells using RIPA buffer containing PMSF (Beyotime, Nantong, China). An equal amount of proteins (100 μg) was separated by 7.5%/12.5% SDS‐PAGE and transferred to a PVDF membrane. Primary polyclonal antibodies targeting TSC1(ab), AMPK Thy (ab133462), AMPK (ab6672), mTOR Ser (ab2486), mTOR (ab142349), Anti‐S6K1 antibody [E175] (ab32359), Anti‐S6K1 (phospho T389) antibody (ab2571), Anti‐EGR2 antibody [EPR4004] (ab108399) were purchased from Abcam (Cambridge, MA, USA). Secondary antibodies were anti‐rabbit or anti‐mouse HRP‐linked and purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Blots were developed using ECL reagent (Millipore, Burlington, MA, USA). An equal amount of protein loading in each lane was confirmed using β‐actin antibody. ImageJ software (NIH, Bethesda, MD, USA) quantified the integrated density of the band.
4
Protein Expression Analysis in 3D Co-Culture
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For western blotting, proteins were extracted from tissues or cultured cells using RIPA buffer containing phenylmethanesulfonylfluoride (PMSF) (Beyotime, Nantong, China). Equal amounts of protein (100 µg) were separated by SDS-PAGE on a 7.5%/12.5% gel and transferred to a PVDF membrane. Primary polyclonal antibodies targeting IκBα(ab7217), IκBα s36 (ab133462), IL-6 (ab6672), TGF-β (ab2486), PAI-1 (ab142349), SMAD3 (ab40584) and p-SMAD3 s425 (ab51177) were purchased from Abcam. The secondary antibodies used were HRP-conjugated anti-rabbit or anti-mouse antibodies and were purchased from Santa Cruz Biotechnology. The blots were developed using ECL reagent (Millipore, MA, USA). An equal amount of loaded protein in each lane was confirmed using a β-actin antibody. ImageJ software was used to quantify the integrated density of the band. ELISA Levels of PAI-1, TGF-β, CCL-17, IL-6 and CCL-22 were measured in the supernatants from cell cultures with a commerical ELISA kit (Thermo Fisher; BMS2033, 88-8350-86, EHCCL17, BMS213-2 and EMCCL22). The experiments were carried out according to the manufacturer's instructions.
3D co-culture system Differentiated THP-1 and 95D cells were cultured in a 3D Petri Dish (Micro-Tissues) (RI, USA) per manufacturer's instructions. Cell culture supernatants were collected daily for seven days. The cells were then collected, washed with PBS, and stained.
3D co-culture system Differentiated THP-1 and 95D cells were cultured in a 3D Petri Dish (Micro-Tissues) (RI, USA) per manufacturer's instructions. Cell culture supernatants were collected daily for seven days. The cells were then collected, washed with PBS, and stained.
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