Ab125938

Manufactured by Abcam

Ab125938 is a laboratory equipment product from Abcam. It is a specialized device designed for scientific research and analysis purposes. The core function of this product is to facilitate specific tasks or processes within a laboratory setting. However, a detailed description of its intended use or features cannot be provided while maintaining an unbiased and factual approach.

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Lab products found in correlation

5 protocols using ab125938

Characterization of Lung Cell Markers

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Cells were washed with MACS flow buffer (MACS, 130-091-222) and permeabilized with BD Cytofix/Cytoperm (BD, 554714) prior to incubation with antibodies. Cells were labeled for antibodies against CD90 (Abcam, ab3105; Abcam, ab124527; Abcam, ab23894; BD, 555595), CD105 (Abcam, ab107595; Abcam, ab2529; Abcam, ab11414; R&D Systems, Fab10971p), Pro-SPC (Bioss, bs 10067R; Abcam, ab40879), CCSP (Abcam, ab171957), Epcam (Abcam, ab71916, Abcam, ab168828; Life Technologies, a15755), and Aqp5 (Abcam, ab78486; Abcam, ab85905) and detected by Alexa Fluor 488 (Abcam, ab150117, ab150077) or fluorescein isothiocyanate (FitC) (Abcam, ab6840) secondary antibodies. Both unstained and isotype controls (Abcam, ab18419; BD, 559320; Abcam, ab125938) were utilized as controls. Human adipose-derived mesenchymal stem cells (AD-MSCs) were obtained from Lonza. Flow Cytometry was performed on the CytoFlex (Beckman Coulter, Indianapolis, IN) and analyzed using FCS Express (De Novo Software, Glendale, CA) or CytExpert ((Beckman Coulter, Indianapolis, IN).

Dinh P.U., Cores J., Hensley M.T., Vandergriff A.C., Tang J., Allen T.A., Caranasos T.G., Adler K.B., Lobo L.J, & Cheng K. (2017). Derivation of therapeutic lung spheroid cells from minimally invasive transbronchial pulmonary biopsies. Respiratory Research, 18, 132.

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Protein Interaction Analysis via Duolink PLA

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To detect protein interactions in cells, a Duolink^® PLA in-situ kit (92101; Sigma–Aldrich Corp.) was used according to the manufacturer’s instructions. The primary antibodies were as follows: a rabbit anti-UBA52 antibody (EPR4547; ab109230; Abcam) and a mouse anti-CDK6 antibody (ab54576; Abcam). The primary antibody rabbit anti-Immunoglobulin G was used as a control (ab125938; Abcam). Images were acquired with a confocal laser microscope using a ×60 oil immersion objective lens. Dots within cells were counted using Duolink ImageTool (OLINK Bioscience, Uppsala, Sweden). Cells were only counted when the whole cell was in the field of vision.

Kobayashi M., Oshima S., Maeyashiki C., Nibe Y., Otsubo K., Matsuzawa Y., Nemoto Y., Nagaishi T., Okamoto R., Tsuchiya K., Nakamura T, & Watanabe M. (2016). The ubiquitin hybrid gene UBA52 regulates ubiquitination of ribosome and sustains embryonic development. Scientific Reports, 6, 36780.

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KRAS and PDE5 Protein Quantification

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Six dishes of Mia PaCa-2 cells were prepared. Renewed the medium with 20 μmol/L compound 36l and 1% DMSO to every 3 dishes respectively and incubated for another 2 h. Harvested and combined the cells, then added 500 μL of IP lysate (Beyotime, containing 1% protease inhibitor) and lysed the cells on ice for 30 min. The prepared Protein A/G beads (Santa Cruz) were added into the cell lysates. Shacked the tubes at 4 °C for 1 h followed by a centrifuge at 4 °C, 12,000 rpm, 15 min. Quantify the supernatant by the BCA method and adjust the total protein concentration to 5 μg/mL. Respectively added KRAS primary antibody (Abcam #ab275876) and IgG (Abcam #ab125938) antibody to 100 μL the above lysates followed by rotating at 4 °C for 1 h. Add the Protein A/G beads 20 μL respectively into the supernatant above, and rotated on the shaker overnight. Centrifuged and collected the precipitate followed by washed it with 500 μL PBS buffer 3 times. Centrifuged and collected the precipitate followed by denaturation in 2 × loading buffer 60 μL at 95 °C for 10 min. Western blotting experiments and gray-scale statistics were conducted to quantify the KRAS and PDEδ protein.

Chen L., Zhang J., Wang X., Li Y., Zhou L., Lu X., Dong G, & Sheng C. (2021). Discovery of novel KRAS‒PDEδ inhibitors with potent activity in patient-derived human pancreatic tumor xenograft models. Acta Pharmaceutica Sinica. B, 12(1), 274-290.

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Quantifying Sox9 Expression in Human VIC Cultures

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Human VIC cultures were seeded on glass slides at a density of 3 × 10⁴ cells per slide. ADGM, DMEM-XAV-939 and ADGM-XAV-939 were added after 72 h in conventional culture. After 14 days of incubation in the different media, the slides were washed twice with DPBS and fixed in 4°C acetone for 10 minutes. Cells were permeabilised with PBS containing 0.2% TritonX100 for 10 minutes and blocked in PBS containing 1%BSA for 30 minutes. Sox9 antibody was diluted 1:1000 in PBS/BSA, added to the cells and incubated for 1 h at RT. As a control, an isotype monoclonal rabbit antibody IgG (abcam, Ab125938) was used at an equivalent concentration. After washing of the slides, a secondary donkey Anti-Rabbit IgG 488 (1:500, abcam, Ab150073) antibody in PBS/BSA was incubated for 1 h at RT. The cells were DAPI stained and washed with PBS twice and then embedded in fluorescent mounting medium (Fluoromount G, Invitrogen). Staining was evaluated and images were acquired using a Zeiss Observer Z.1 Apotome fluorescence microscope. Fluorescent nuclei were counted in relation to DAPI-stained nuclei and the percentages of Sox9-positive nuclei were evaluated.

Dittfeld C., Reimann G., Mieting A., Büttner P., Jannasch A., Plötze K., Steiner G., Tugtekin S.M, & Matschke K. (2018). Treatment with XAV-939 prevents in vitro calcification of human valvular interstitial cells. PLoS ONE, 13(12), e0208774.

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Quantifying CCR7 Expression in Synovial Membrane

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CCR7 expression in human synovial membrane was examined by routine immunohistochemical (IHC) staining using a monoclonal anti-human CCR7 antibody (ab65851, Abcam, Cambridge MA). Staining specificity was confirmed by utilizing an isotype-matched immunoglobulin control (ab125938, Abcam), and was quantified on 10× images (NIS-Elements AR, Nikon Instruments Inc., Melville, NY). Image analysis involved defining five regions of interest (ROIs, each 2.5 × 10 μm² including lining and sub-lining) on a single section per patient. Red thresholds were set uniformly to highlight positively stained areas, and background was set by comparison to isotype matched controls. Area positively stained in each ROI was summed and reported as a fraction of total area analyzed (1.25 × 106 μm2).

Sambamurthy N., Nguyen V., Smalley R., Xiao R., Hankenson K., Gan J., Miller R.E., Malfait A.M., Dodge G.R, & Scanzello C.R. (2017). Chemokine Receptor-7 (CCR7) Deficiency Leads to Delayed Development of Joint Damage and Functional Deficits in a Murine Model of Osteoarthritis. Journal of orthopaedic research : official publication of the Orthopaedic Research Society, 36(3), 864-875.

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