polyclonal rabbit anti trpv4  (Alomone Labs)

Journal: The Journal of Physiological Sciences : JPS

Article Title: Calcium response via CRAC channels in human synovial cells induced by shear stress in rheumatoid arthritis

doi: 10.1016/j.jphyss.2025.100013

Figure Lengend Snippet: Immunofluorescence staining, differential interference contrast (DIC) microscopy images, and immunofluorescence intensity of the normal and RA FLS cells, Immunofluorescence staining with anti-Ca 2+ release-activated channel regulator 2 A (CRACR2A) antibody (green, Alexa) and DIC images of the normal (A) and RA (B) FLS cells. Immunofluorescence staining with the anti-transient receptor potential vanilloid 4 (TRPV4) antibody (red, Alexa) and DIC images of the normal (C) and RA (D) FLS cells. All fluorescence and DIC images were obtained at 20 × magnification using an Olympus FV1000 Confocal Laser Microscope. Scale bar = 100 µm. The mean immunofluorescence intensity of the anti-CRACR2A antibody staining was significantly lower in the normal FLS cells (4.08 ± 0.8 AU) than in the RA FLS cells (20.3 ± 6.9 AU) (p < 0.01). The mean immunofluorescence intensities produced by the anti-TRPV4 antibody staining were not significantly different between the normal (30.3 ± 6.2 AU) and RA (29.7 ± 6.2 AU) FLS cells (E). n.s.: not significant, * *: p < 0.01 , compared between the normal and RA FLS cells.

Article Snippet: After fixation, the cultures were washed twice with PBS and blocked with 0.2 % bovine serum in PBS for 30 min. Primary antibodies were diluted (1:250 for rabbit polyclonal anti-CRACR2A antibody [66787–1-IG, Proteintech, IL, USA] and 1:250 for rabbit polyclonal anti-TRPV4 antibody [ab191580, Alomone Labs, Jerusalem, Israel]) and incubated for 1 h at 37.5 °C.

Techniques: Immunofluorescence, Staining, Microscopy, Fluorescence, Produced

Journal: The Journal of Physiological Sciences : JPS

Article Title: Calcium response via CRAC channels in human synovial cells induced by shear stress in rheumatoid arthritis

doi: 10.1016/j.jphyss.2025.100013

Figure Lengend Snippet: Expression levels of ORAI1, stromal interaction molecule 1 (STIM1), CRACR2A, and TRPV4 in the normal and RA FLS cells Western blotting of ORAI1, STIM1, CRACR2A, and TRPV4 expression in the normal and RA FLS cells. (A) The internal controls and representative western blots are shown. Subsequently, 10 or 30 μg of protein per lane was loaded onto the gel. Next, the membranes were probed with specific antibodies against Orai1, STIM1, CRACR2A, or TRPV4 with appropriate secondary antibodies. Protein bands were detected using C-DiGit (LI-COR#3600–00), and the results were analyzed using the ImageJ software (B). The protein amounts were normalized to β-actin amounts to indicate the protein expression ratios. * * and † † represent significant differences from normal FLS, respectively, at p < 0.01.

Article Snippet: After fixation, the cultures were washed twice with PBS and blocked with 0.2 % bovine serum in PBS for 30 min. Primary antibodies were diluted (1:250 for rabbit polyclonal anti-CRACR2A antibody [66787–1-IG, Proteintech, IL, USA] and 1:250 for rabbit polyclonal anti-TRPV4 antibody [ab191580, Alomone Labs, Jerusalem, Israel]) and incubated for 1 h at 37.5 °C.

Techniques: Expressing, Western Blot, Software

Journal: The Journal of Physiological Sciences : JPS

Article Title: Calcium response via CRAC channels in human synovial cells induced by shear stress in rheumatoid arthritis

doi: 10.1016/j.jphyss.2025.100013

Figure Lengend Snippet: Ca 2+ influx from the CRAC and TRPV4 of synovial cells in response to SS, When the Ca 2+ in the endoplasmic reticulum was depleted by SS, CRACR2A directly interacted with the cytoplasmic regions of ORAI1 and STIM1 to form a ternary complex, resulting in the intracellular Ca 2+ uptake from extracellular sources. In the RA FLS cells, STIM1 and CRACR2A were overexpressed compared with the expression in the normal FLS cells, and the regulation of intracellular Ca 2+ concentration significantly depended on CRAC. SS also induced Ca 2+ uptake from the extracellular into the intracellular site via TRPV4 and/or other mechanosensitive Ca 2+ channels. However, there were no significant difference in the expression level of TRPV4 between the normal and RA FLS cells, indicating that TRPV4 exerted the same influence on the regulation of intracellular Ca 2+ concentration in both FLS cell types.

Article Snippet: After fixation, the cultures were washed twice with PBS and blocked with 0.2 % bovine serum in PBS for 30 min. Primary antibodies were diluted (1:250 for rabbit polyclonal anti-CRACR2A antibody [66787–1-IG, Proteintech, IL, USA] and 1:250 for rabbit polyclonal anti-TRPV4 antibody [ab191580, Alomone Labs, Jerusalem, Israel]) and incubated for 1 h at 37.5 °C.

Techniques: Expressing, Concentration Assay