Ab92360

Manufactured by Abcam

Sourced in United States

Ab92360 is a laboratory equipment product designed for use in scientific research. It serves as a core function in various experimental procedures. The detailed specifications and intended use of this product are not available for an unbiased and factual description. Further information may be obtained by contacting the manufacturer.

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Lab products found in correlation

Ab181544, by Abcam (2 mentions) Ab137096, by Abcam (2 mentions) Hc201 01, by Transgene (2 mentions) Protease inhibitor, by Roche (2 mentions) Ventana discovery ultra, by Roche (1 mentions) Nanozoomer slide scanner, by Hamamatsu Photonics (1 mentions) Sds polyacrylamide gels, by National Diagnostics (1 mentions)

4 protocols using ab92360

Immunohistochemical Analysis of RanGAP1 in ALS

Cited 1 time

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Tissue sections were cut from blocks of formalin-fixed paraffin embedded ALS tissue, obtained from Dr. John Ravits’ bank collection (control patient #83 and #65). The spinal cord sections of patients harboring TDP-43 mutations (Pt #284 and #203) as well as control #150 were obtained from the MRC London Neurodegenerative Diseases Brain Bank (King’s College London) (see table below). Seven µm-thick tissue sections were stained with an antibody against RanGAP1 (ab92360) from Abcam (Cambridge, MA, USA). Slides were stained on a Ventana Discovery Ultra (Ventana Medical Systems, Tucson, AZ, USA). Antigen retrieval was performed using Tris–EDTA based cell conditioning solution (CC1) for 40 min at 95 °C. The primary antibody was incubated on the sections at 1:300 dilution for 32 min at 37 °C followed by UltraMap (Ventana) and DAB detection. Slides were rinsed, dehydrated through alcohol and xylene, and coverslipped. Imaging was performed on Nanozoomer slide scanner (Hamamatsu) at the UCSD microscopy core.

Patient #	Age at death (years)	Sex	Diagnosis	Genotype	Post-mortem delay + interval (h)	Reference
65	82	M	NORMAL	N/A	4*
83	63	F	NORMAL	N/A	4*
A150/01	40	M	NORMAL	N/A	40
A284/13	57	M	ALS	TARDBP M337V	74	[60 (link)]
A203/12	23	F	ALS	TARDBP Y374X + FUS P525L	125	[32 (link)]

* Post-mortem interval only (post-mortem delay not available)

Ditsworth D., Maldonado M., McAlonis-Downes M., Sun S., Seelman A., Drenner K., Arnold E., Ling S.C., Pizzo D., Ravits J., Cleveland D.W, & Da Cruz S. (2017). Mutant TDP-43 within motor neurons drives disease onset but not progression in amyotrophic lateral sclerosis. Acta Neuropathologica, 133(6), 907-922.

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Western Blot Protein Analysis

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Cells were lysed in RIPA buffer (50 mM Tris-HCl at pH 7.4, 150 mM NaCl, 0.1% SDS, 1% NP-40, 0.25% sodium deoxycholate, and 1 mM dithiothreitol) supplemented with protease inhibitors (Roche, Basel, Switzerland) and incubated for 30 min on ice. After centrifugation at 18,800 g for 10 min, the protein concentration in the supernatant was determined using the BCA method. Equal amounts of protein were separated by sodium dodecyl sulfate (SDS)-sulfate-polyacrylamide gel electrophoresis in Tris/glycine/SDS running buffer. The separated proteins were transferred onto a polyvinylidene fluoride membrane in Tris-glycine transfer buffer that was blocked with 3% Tris-buffered saline with Tween 20 (BSA-TBST) for 1 h at room temperature. The membranes were probed with primary RanGAP1 antibody (ab92360, Abcam) at a 1:500 dilution and HBG2 (ab137096, Abcam), GATA1 (ab181544, Abcam), and β-actin antibodies (HC201-01, TransGen Biotech, Beijing, China) at a 1:1,000 dilution in 3% BSA-TBST at 4°C overnight. The membranes were subsequently rinsed with TBST three times for 10 min each and incubated with a secondary antibody for 1 h at room temperature. Finally, the membranes were rinsed three times with TBST solution and imaged using an Image Quant ECL machine (Tanon 5200, Tanon, Shanghai, China).

Xiao R., Zhang L., Xin Z., Zhu J., Zhang Q., Zheng G., Chu S., Wu J., Zhang L., Wan Y., Chen X., Yuan W., Zhang Z., Zhu X, & Fang X. (2024). Disruption of mitochondrial energy metabolism is a putative pathogenesis of Diamond-Blackfan anemia. iScience, 27(3), 109172.

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Silencing βarrestin2 in HEK-293 Cells

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HEK-293 or HEK-293 T cells stably expressing the β 2 AR were transfected with either non-targeting control siRNA or siRNA targeting βarr2 purchased from Dharmacon GE Healthcare Life Sciences as described previously [5] . Early passage cells on 6-well dishes, at a confluence of 40-50% were transfected with 3.5 μg siRNA using Lipofectamine 2000™ in serum-free media. After 4 h, complete media was added to the transfected cells, and then grown for 48 h at 37 °C before conducting assays. Cells were serum starved for 1 h prior to stimulation with 1 μM isoproterenol for 20 min. After stimulation, cells were solubilized by adding 2×-SDS-sample buffer, followed by disruption by sonication. Equal amount of cell lysates were resolved on 10% SDSpolyacrylamide gels (ProtoGel, National Diagnostics). RanGAP1, βarrestins and GAPDH were detected by immunoblotting with rabbit monoclonal anti-RanGAP1antibody (Abcam ab92360, 1:1000), antiβarrestin (A1CT, 1:3000) and rabbit monoclonal anti-GAPDH (HRP conjugate, CST 3683, 1:1000) respectively.

Nagi K., Kaur S., Bai Y, & Shenoy S.K. (2020). In-frame fusion of SUMO1 enhances βarrestin2's association with activated GPCRs as well as with nuclear pore complexes. Cellular signalling, 75.

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Western Blot Analysis of Cellular Proteins

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Cells were lysed in RIPA buffer (50 mM Tris-HCl at pH 7.4, 150 mM NaCl, 0.1% SDS, 1% NP-40, 0.25% sodium deoxycholate, 1 mM dithiothreitol) supplemented with protease inhibitors (Roche, Basel, Switzerland) and incubated for 30 min on ice. After centrifugation at 18,800 ×g for 10 min, the protein concentration in the supernatant was determined. Equal amounts of protein were separated using sodium dodecyl sulfate (SDS)polyacrylamide gel electrophoresis in a Tris/glycine/SDS running buffer. The separated proteins were transferred onto a polyvinylidene fluoride membrane in Tris/glycine transfer buffer, which was then blocked with 3% BSA-TBST (Tris buffered saline with Tween 20) for 1 h at room temperature. The membranes were probed with primary RanGAP1 mouse monoclonal antibody (ab92360, Abcam) at a 1:500 dilution and HBG2 (ab137096, Abcam), GATA1 (ab181544, Abcam), and β-actin mouse monoclonal antibodies (HC201-01, TransGen Biotech, Beijing, China) at a 1:1000 dilution in 3% BSA-TBST at 4℃ overnight. The membranes were subsequently rinsed with TBST thrice for 10 min each and incubated with secondary antibody for 1 h at room temperature. Finally, the membranes were rinsed with TBST solution thrice and imaged using an Image Quant ECL machine (Tanon 5200, Tanon, Shanghai, China).

Xiao R., Zhang L., Xin Z., Zhu J., Zhang Q., Chu S., Wu J., Zhang L., Wan Y., Chen X., Yuan W., Zhang Z., Zhu X., & Fang X. (2022). Oxidative phosphorylation pathway disruption is an alternative pathological mechanism leading to Diamond-Blackfan anemia.

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