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Monoclonal rabbit antibody

Manufactured by Cell Signaling Technology
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Monoclonal rabbit antibodies are laboratory reagents used to detect and measure specific proteins or molecules in biological samples. They are produced by cloning a single B lymphocyte, ensuring a consistent and uniform antibody targeting a specific epitope. These antibodies provide a reliable and specific tool for various research and diagnostic applications.

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Lab products found in correlation

Zen software, by Zeiss (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions) Lsm 800, by Zeiss (1 mentions) Monoclonal mouse antibody, by Abcam (1 mentions) Target retrieval solution ph 6, by Agilent Technologies (1 mentions) Target retrieval solution ph 9, by Agilent Technologies (1 mentions) Envision kit, by Agilent Technologies (1 mentions) A0545, by Merck Group (1 mentions) Tgx gel, by Bio-Rad (1 mentions) Nitrocellulose membrane, by Cytiva (1 mentions) Polyvinylidene fluoride (pvdf), by Merck Group (1 mentions) Bca protein assay, by Beyotime (1 mentions) Wnt3a, by Abcam (1 mentions) Radio immunoprecipitation assay ripa buffer, by Beyotime (1 mentions) Phosphatase inhibitor, by Thermo Fisher Scientific (1 mentions) Hyclone fbs, by Thermo Fisher Scientific (1 mentions) Odyssey imaging system, by LI COR (1 mentions) Phospho 4ebp1, by Cell Signaling Technology (1 mentions) Ripa buffer, by Merck Group (1 mentions) Protease inhibitor cocktail, by Merck Group (1 mentions)

Close spelling variants of this product

Rabbit monoclonal anti-PTEN antibody Anti-phosphotyrosine rabbit monoclonal antibody (P-Tyr-1000) beads Primary rabbit monoclonal anti-GAPDH antibody Beta-actin rabbit monoclonal antibody Rabbit monoclonal antibody against HA GSK-3β Rabbit Monoclonal Antibody Rabbit monoclonal antibody against cleaved caspase-3 Rabbit anti-p53 monoclonal antibody Rabbit monoclonal F4/80 antibody Rabbit monoclonal anti-GAPDH antibody

8 protocols using monoclonal rabbit antibody

1

Western Blot Analysis of SH-SY5Y Cells

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For Western blot analysis, SH-SY5Y cells were transfected with the miR-129-5p mimic as described above. After 48 h, the cells were harvested, lysed in 2x sample buffer (130 mM Tris/HCl, 6% [v/v] SDS, 10% [v/v] 3-mercapto-1,2-propandiol, and 10% [v/v] glycerol) and sonicated three times for three seconds each. Total protein extract (10 µg) was electrophoresed in a 4-15% TGX gel (Bio-Rad Laboratories, Inc., Hercules, California, USA). The proteins were electroblotted onto nitrocellulose membranes (Whatman, GE Healthcare, Freiburg, Germany). Unspecific antibody binding was blocked by preincubation of the nitrocellulose membrane in 5% TBS milk with 0.1% Tween 20 for 30 min. SNCA was detected by a polyclonal rabbit antibody (#2642), COMT by a monoclonal rabbit antibody (#14368), CLOCK by a monoclonal rabbit antibody (#5157), and AKT3 by a monoclonal rabbit antibody (#14982), all of which were purchased from Cell Signaling Technology (Danvers, MA, USA). α-Tubulin served as an endogenous control and was detected by a monoclonal rabbit antibody (#2125 Cell Signaling Technology, Danvers, MA, USA). A secondary anti-rabbit antibody was purchased from Sigma‒Aldrich (A0545; Sigma Aldrich, Munich, Germany).
Hart M., Kern F., Fecher-Trost C., Krammes L., Aparicio E., Engel A., Hirsch P., Wagner V., Keller V., Schmartz G.P., Rheinheimer S., Diener C., Fischer U., Mayer J., Meyer M.R., Flockerzi V., Keller A, & Meese E. (2024). Experimental capture of miRNA targetomes: disease-specific 3′UTR library-based miRNA targetomics for Parkinson’s disease. Experimental & Molecular Medicine, 56(4), 935-945.
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2

Coronary Plaque APOL1 and Apoptosis

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Coronary arteries were stained against APOL1 (Sigma, St. Louis, MO) and CD68 (Dako, Carpinteria, CA). In addition, a subset of 14 stable plaques (including plaques from 5 carriers of the reference allele, 5 carriers of one, and 4 carriers of two APOL1 risk alleles) and 16 ruptured coronary plaques (from 6 carriers of the reference allele, 5 carriers of one, and 5 carriers of two APOL1 risk alleles) was stained against apolipoprotein A1 (APOA1), detected by a polyclonal APOA1 rabbit antibody (Sigma, St. Louis, MO), and cleaved caspase 3, identified by a monoclonal rabbit antibody (Cell Signaling Technology). Nuclear counterstain was performed with DAPI (Invitrogen). Images were obtained with a confocal laser scanning microscope (LSM 800, Zeiss, Oberkochen, Germany), and staining quantification was performed using Zen software (Zeiss, Oberkochen, Germany), assessing the APOL1-, CD68, APOA1-, and cleaved caspase 3-positive areas within the plaque area, defined as the area between the IEL and the lumen.
Cornelissen A., Fuller D.T., Fernandez R., Zhao X., Kutys R., Binns-Roemer E., Delsante M., Sakamoto A., Paek K.H., Sato Y., Kawakami R., Mori M., Kawai K., Yoshida T., Latt K.Z., Miller C.L., de Vries P.S., Kolodgie F.D., Virmani R., Shin M.K., Hoek M., Heymann J., Kopp J.B., Rosenberg A.Z., Davis H.R., Guo L, & Finn A.V. (2021). APOL1 Genetic Variants are Associated with Increased Risk of Coronary Atherosclerotic Plaque Rupture in African Americans. Arteriosclerosis, thrombosis, and vascular biology, 41(7), 2201-2214.
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3

Immunohistochemical Staining Protocol

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For the detection of GATA3, a monoclonal rabbit antibody (Cell Signalling Technologies) was used. FLG was stained with a monoclonal mouse antibody purchased from Abcam and for FLG2detection, a polyclonal rabbit antibody (Sigma-Aldrich) was used. For all three targets the Envision+ kits from Dako (Hamburg, Germany) were used (FLG: HRP.Mouse (AEC+); GATA3: HRP.Rabbit (DAB+); FLG2: HRP.Rabbit AEC+)). The detection of Lucifer Yellow was performed with a polyclonal rabbit antibody (bought from ThermoScientific) in combination with HRP-Green Solution Set (42 life sciences GmbH & Co. KG, Bremerhaven, Germany). In all experiments the corresponding isotype control was added as a control. For the staining of FLG, GATA3, and Lucifer Yellow the “Target Retrieval Solution, ph9” and for FLG2 the “Target Retrieval Solution, ph6” (both Dako) was used. An Axio Scan.Z1 (Zeiss, Jena, Germany) was used for digitalization of the immunohistological specimens depicted in this study. Quantification of the staining intensity was done by using the software cellSense Dimension with Count & Measure Solution (Olympus Deutschland GmbH, Hamburg, Germany).
Zeitvogel J., Jokmin N., Rieker S., Klug I., Brandenberger C, & Werfel T. (2017). GATA3 regulates FLG and FLG2 expression in human primary keratinocytes. Scientific Reports, 7, 11847.
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4

Immunoblotting with Phospho-Specific Antibodies

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The following antibodies were used: monoclonal rabbit antibody against phosphorylated ezrin/radixin/moesin (pERM; product # 3149; Cell Signaling, Danvers, MA, USA), monoclonal mouse antibody clone PY99 against phosphotyrosine (pY; Santa Cruz Biotechnology Inc., Santa Cruz, CA, USA), and polyclonal rabbit antibody against histone H3 phosphorylated at Ser10 (H3-P; product # 06–570; Merck Millipore, Billerica, MA, USA). Cross-reactivity of anti-pERM with honeybee moesin has been demonstrated previously [35 (link)].
Klose S.P., Rolke D, & Baumann O. (2017). Morphogenesis of honeybee hypopharyngeal gland during pupal development. Frontiers in Zoology, 14, 22.
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5

Western Blot Analysis of Protein Levels

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RCECs were lysed with Radio Immunoprecipitation Assay (RIPA) buffer (Biyuntian, Shanghai, China). The cells were centrifuged at 14,000g for 20 min at 4 °C after ultrasonic shock by ice lysis, and the protein concentration was determined using the BCA protein assay (Biyuntian, Shanghai, China). Proteins were separated by dodecyl sulfate and sodium salt-polyacrylamide gel (SDS-PAGE) electrophoresis. Proteins of 30–50 µg were separated by 12.5% SDS–polyacrylamide gel electrophoresis. Total protein levels were determined with Wnt3a (abcam, ab219412), PDLIM1 (abcam, ab129015) and Actin (MDL, MDL11027) monoclonal rabbit antibodies (Cell Signaling Technology, USA). All antibodies were diluted 1:1000. After the application of the primary antibody, the PVDF (Millipore, Germany) was washed three times and then incubated with the secondary antibody, which was coupled with horseradish peroxidase and incubated for 2 h at 37 °C. The luminescence image analyzer Image J software (National Institutes of Health, America) was used to quantify the Western blot bands, and the optical densities of target genes were normalized by ACTIN.
Xie P., You Q., Zhu J., Xie W., Wei P., Zhu S., Du Y, & Gao X. (2023). PDLIM1 inhibits cell migration and invasion in diabetic retinopathy via negatively regulating Wnt3a. Scientific Reports, 13, 5820.
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6

Immunoblotting Analysis of AMPK Signaling

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Unless otherwise indicated, all immunoblots depict relative protein from whole B cell extracts derived from splenocytes of Rosa26-CreERT2 mice (Prkaa1 +/+ or Prkaa1 f/f) followed by a 2-day culture with LPS, BAFF, and 4-OHT at 5 × 106 per mL. For immunoblotting for AMPK targets after glucose starvation, 2-day LPS blasts were washed and reseeded in glucose-free RPMI (Invitrogen) supplemented with 10% dialyzed HyClone FBS (Thermo Scientific) for specified amount of time. Cells were washed twice in cold PBS and lysed in RIPA buffer (Sigma catalog # 0278) in the presence of phosphatase inhibitor (Thermo Scientific) and a protease inhibitor cocktail (Sigma). Twenty to one hundred micrograms of cell lysates were resolved by SDS-PAGE, transferred to PVDF membranes, and immunoblotted for phospho-AMPKα1 (T172), AMPKα, phospho-S6 (S235/236), total S6, phospho-ACC (S79), total ACC, phospho-ULK1 (S317), total ULK1, phospho-4E-BP1, total 4E-BP1, and/or LC3 using monoclonal rabbit antibodies purchased from Cell Signaling Technology (Danvers, MA). Actin (Santa Cruz Biotechnology, Dallas, TX) was detected on all blots as a loading control. Immunoblots were visualized using Odyssey Imaging system (Li-Cor) after incubation with secondary reagents, anti-rabbit IgG-680, or anti-mouse IgG-800 (Invitrogen).
Brookens S.K., Cho S.H., Basso P.J, & Boothby M.R. (2020). AMPKα1 in B cells dampens primary antibody responses yet promotes mitochondrial homeostasis and persistence of B cell memory. Journal of immunology (Baltimore, Md. : 1950), 205(11), 3011-3022.
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7

Quantitative Analysis of miRNA Targets

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Mimics and inhibitors of miR-497 were transfected into cells, and RNA and protein were extracted from transfected cells after 48 h. MiRNA expression levels were determined with the S-Poly (T) method (16). Total RNA (1 µg) was used for cDNA synthesis using a HiScript ® QRT SuperMix (+ gDNA wiper) (Nanjing, China) kit for qPCR (R123-1, Vazyme, Nanjing, China). The primers were designed based on the sequences of bovine PPARG, ACLY, CD36, ELOVL6, HSL, and GAPDH in NCBI (Table 2), and real-time PCR results were analyzed using the 2−ΔΔCt model.
SDS-PAGE was used to isolate proteins from cells, and they were transferred nitrocellulose membrane (Millipore, Billerica, MA, USA). Rabbit monoclonal (# 8579, Cell Signaling Technology, Shanghai, China) and monoclonal rabbit antibodies (# 8457, Cell Signaling Technology, Shanghai, China) were used, and image collection and analysis were conducted.
Chu S., Yang Y., Nazar M., Chen Z, & Yang Z. (2023). miR-497 Regulates LATS1 through the PPARG Pathway to Participate in Fatty Acid Synthesis in Bovine Mammary Epithelial Cells. Genes, 14(6), 1224.
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8

Protein Expression and Localization

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The primary antibodies, p-p53, p-chk2, chk2, γ-h2ax, p-γ-h2ax, monoclonal rabbit antibodies, P53 and GAPDH mouse antibodies were got from cell signalling technology (Danvers, MA). Rabbit anti-human antibodies against p21, Bax, bcl2, cyclin B, and caspase 3 were obtained from proteolytic enzyme technology (Wuhan, China). Secondary antibody was purchased from proteineach technology (Wuhan, China). Red fluorescent antibody, Cy3 goat anti-rabbit and anti-mouse secondary antibody were purchased from Invitrogen (Carlsbad, CA). The matrix gel (356234) was obtained from Corning (Corning, NY).
Sun L., Wu C., Ming J., Nie X., Guo E., Zhang W, & Hu G. (2020). Riluzole Enhances the Response of Human Nasopharyngeal Carcinoma Cells to Ionizing Radiation via ATM/P53 Signalling Pathway. Journal of Cancer, 11(11), 3089-3098.
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