Protein samples from MEFs or neonatal tissues were separated with SDS-PAGE and transferred to PVDF or nitrocellulose membranes, respectively. Membranes were blocked with 5% skim milk, and blotted with anti-LTBP-1 polyclonal antibody (Ab39, a generous gift from Drs. K Miyazono and C–H Heldin [37 (link)], or ab78294 (Abcam)) as a primary antibody and horseradish peroxidase-conjugated anti-Rabbit IgG antibody (GE Healthcare Life Science) as a secondary antibody. Chemiluminescent reations were performed with either ECL Western Blotting Substrate (Thermo Scientific) or Western Lightning Chemiluminescence Plus-ECL (Perkin Elmer), and signals were detected with X-ray films or ImageQuant LAS 4000 System (GE Healthcare Life Science).
Ab78294
Manufactured by Abcam
Sourced in United Kingdom
Ab78294 is a lab equipment product used for scientific research and analysis. It is a high-quality tool designed to assist researchers in their laboratory work. The core function of this product is to provide a reliable and efficient solution for the task it is intended to perform. However, without more specific details about the product, I cannot provide a detailed description while maintaining an unbiased and factual approach. Therefore, the description for this product is not available.
Automatically generated - may contain errors
Lab products found in correlation
Imagequant las 4000 system, by GE Healthcare (1 mentions)
Horseradish peroxidase conjugated anti rabbit igg antibody, by GE Healthcare (1 mentions)
Ecl western blotting substrate, by Thermo Fisher Scientific (1 mentions)
Polyvinylidene fluoride (pvdf), by Merck Group (1 mentions)
Ibright 1500, by Thermo Fisher Scientific (1 mentions)
Ab76013, by Abcam (1 mentions)
Ab205394, by Abcam (1 mentions)
D9564, by Merck Group (1 mentions)
A2228, by Merck Group (1 mentions)
Nbp1 91258, by Novus Biologicals (1 mentions)
Ab54208, by Abcam (1 mentions)
Mab5871, by R&D Systems (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions)
F3165, by Merck Group (1 mentions)
β actin, by Merck Group (1 mentions)
3 protocols using ab78294
1
Western Blot Analysis of LTBP-1
2
Western Blot Analysis of Protein Expression
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For the analysis of total proteins, lung tissues or PASMCs were lysed by RIPA buffer with a cocktail and PMSF and homogenated by tissue homogenator (Wuhan Service Biotechnology). The lysates were centrifuged for 15 min at 13000 g, 4℃. The concentration of total protein was determined by the BCA assay kit (Beijing Biyuntian Biotechnology). The lysates were eluted with 2*SDS buffer and boiled for 10 min at 100℃. The samples were loaded and separated on 10% SDS/PAGE gel, and then transferred onto polyvinylidene difluoride membranes (PVDF) (Millipore, USA). The membranes were blocked in 5% non-fat milk and then incubated with specific primary antibodies (anti β-actin Mouse antibody [GB12001, Wuhan Servicebio Technology, China], and anti-LTBP1 antibody [ab78294, Abcam, UK]) at a dilution of 1:1000 at 4 °C overnight. After being incubated with HRP-conjugated secondary antibodies, the blots were visualized using the ECL system (New Cell and Molecular Biotech Co, Ltd, China), captured by iBright 1500 (Invitrogen by Thermo Fisher Scientific) and analyzed by Image J software.
3
Immunoblotting and Immunofluorescence Analysis
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Immunoblotting was performed as described previously [44 (link)]. Equal amounts of tissue lysates were used for immunoblotting. Blots were incubated with specific antibodies to BMP-1 and latent TGFβ1-binding protein-1 (LTBP-1) (ab205394 and ab78294; Abcam), Flag (F3165; Sigma-Aldrich), ALK5 (MAB5871; R&D Systems), Col3a1 and Fn1 (NB600 and NBP1-91258; Novus Biologicals). β-Actin (A2228; Sigma-Aldrich) was used as a loading control. Immunofluorescence was performed as described in detail previously [44 (link)]. We used specific antibodies to CD34 (553731; BD Bioscience or ab54208; Abcam), Nkx2.1 (ab76013; Abcam) and VE-cadherin (562243; BD Biosciences). The nuclei were stained with 4′,6-diamidino-2-phenylindole (D9564; Sigma-Aldrich).
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