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Ab8245

Manufactured by Merck Group
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Ab8245 is a laboratory equipment product manufactured by Merck Group. It is designed for general laboratory use. The core function of Ab8245 is to perform precise measurements and analyses within a controlled laboratory environment.

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Lab products found in correlation

Ab134936, by Abcam (2 mentions) Ab130769, by Abcam (2 mentions) Ab16478, by Abcam (2 mentions) Casein blocking buffer, by Merck Group (2 mentions) C6219, by Merck Group (2 mentions) Alexa fluor 680, by Thermo Fisher Scientific (2 mentions) Irdye 800, by Rockland Immunochemicals (2 mentions) Odyssey infrared imaging system, by LI COR (2 mentions) Nitrocellulose membrane, by Bio-Rad (2 mentions) Ab1791, by Abcam (2 mentions) Ab15093, by Abcam (1 mentions) Ab97672, by Abcam (1 mentions) T8328, by Merck Group (1 mentions) F3165, by Merck Group (1 mentions) Ab8295, by Abcam (1 mentions) Ab14348, by Abcam (1 mentions) Anti orai1, by Merck Group (1 mentions) Dc protein assay, by Bio-Rad (1 mentions) Anti gapdh, by Abcam (1 mentions) Ab54615, by Abcam (1 mentions)

11 protocols using ab8245

1

Meiotic Proteins Immunofluorescence and Western Blot

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The following antibodies were purchased: anti-SYCP3 (Abcam, ab97672, IF: 1:100), anti-SYCP3 (Abcam, ab15093, IF: 1:100), anti-SYCP1 (Abcam, ab15090, IF: 1:100), anti-γH2AX (MerckMillipore, 16-202A, IF: 1:800), anti-TNP2 (Santa Cruz, sc-393843, IF: 1:50), anti-PRM2 (Briar Patch Biosciences, Hup 2B, IF: 1:300), anti-H4K8ac (ABclonal, A7258, IF: 1:500), anti-ELFN2 (Novus Biologicals, 90569, WB: 1: 1000), anti-H3 (Abcam, ab1791, WB: 1: 2000), anti-GAPDH (Abcam, ab8245, WB: 1: 10000), anti-TUBLIN (Sigma, T8328, WB: 1: 2000), anti-FLAG (Sigma, F3165, IP: 10 µg, WB: 1:2000). For immunofluorescence studies, the following secondary antibodies were used: anti-rabbit (Jackson ImmunoResearch 488, 711225152, IF: 1:350), anti-rabbit (Jackson ImmunoResearch 594, 711585152, IF: 1:350), anti-mouse (Jackson ImmunoResearch 488, 715545150, IF: 1:350). For Western blot analyses, the following secondary antibodies conjugated to Horse Radish Peroxidase were used: anti-rabbit IgG HRP-linked (ABclonal, AS014, WB: 1:5000), anti-mouse IgG HRP-linked (ABclonal, AS003, WB: 1:5000).
Tan H., Wang W., Zhou C., Wang Y., Zhang S., Yang P., Guo R., Chen W., Zhang J., Ye L., Cui Y., Ni T, & Zheng K. (2023). Single-cell RNA-seq uncovers dynamic processes orchestrated by RNA-binding protein DDX43 in chromatin remodeling during spermiogenesis. Nature Communications, 14, 2499.
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2

Western Blot Analysis of Alzheimer's Biomarkers

Cited 2 times
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Brain homogenates (frontal cortex) in urea (50 μg) were mixed with Laemmli sample buffer and resolved by SDS-PAGE before an overnight wet transfer to nitrocellulose membranes (BioRad) as previously reported (Seyfried et al., 2017 (link)). Membranes were blocked with casein blocking buffer (Sigma B6429) and probed with primary polyclonal antibodies for Anti-PLCD1 antibody (Abcam #ab134936), anti-HEPACAM (Abcam #ab130769), anti-PADI2 antibody (Abcam #ab16478), anti-GAPDH antibody (Abcam #ab8245), and anti-Connexin-43 (Sigma #C6219) overnight at 4°C. Membranes were incubated with secondary antibodies conjugated to Alexa Fluor 680 (Invitrogen) or IRDye800 (Rockland) fluorophores for one hour at room temperature. Images were captured using an Odyssey Infrared Imaging System (Li-Cor Biosciences). Protein densitometry for relative quantification was performed using ImageJ open source software. For GJA1, the higher molecular weight band reflective of ubiquitination (Abreha et al., 2018 (link)) was analyzed separately from the lower molecular weight band. Densitometry was compared between AD and control using the Student’s t test (Student, 1908 ).
Swarup V., Chang T.S., Duong D.M., Dammer E.B., Dai J., Lah J.J., Johnson E.C., Seyfried N.T., Levey A.I, & Geschwind D.H. (2020). Identification of Conserved Proteomic Networks in Neurodegenerative Dementia. Cell reports, 31(12), 107807.
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3

Western Blot Protein Analysis

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Cellular and tissue proteins were extracted with lysis buffer, which contained 1% (vol/vol) Nonidet P-40, 150 mmol/L NaCl, 20 mmol/L Tris-HCl, pH 8.0, with the addition of Roche protease inhibitor cocktail tablets (Sigma–Aldrich, St. Louis, MO, USA, 04693132001). The protein concentrations were determined with the DC Protein Assay (Bio-Rad, Hercules, CA, USA, 500-0002). The proteins were separated in 12% SDS-PAGE and transferred to PVDF membranes. After blocking with 5% BSA in TBS for 2 h at room temperature, the membranes were incubated with primary antibodies at 4°C overnight. The primary antibody was anti-GAPDH (1:5000, Abcam, Cambridge, UK, ab8245), anti-Orai1 (1:1000, Sigma–Aldrich, St. Louis, MO, USA, O8264), anti-STIM1 (1:1000, Cell Signaling, Danvers, MA, USA 5668S), anti-ANF (1:3000, Abcam, ab14348), or anti-cTnT (1:3000, Abcam, ab8295). Immunodetection was accomplished using horseradish peroxidase-conjugated secondary antibody, followed by analysis via the ECL detection system. The intensity of immunoblot bands was quantified using ImageJ Software.
Zheng C., Lo C.Y., Meng Z., Li Z., Zhong M., Zhang P., Lu J., Yang Z., Yan F., Zhang Y., Huang Y, & Yao X. (2017). Gastrodin Inhibits Store-Operated Ca2+ Entry and Alleviates Cardiac Hypertrophy. Frontiers in Pharmacology, 8, 222.
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4

Western Blot Analysis of FAAH Proteins

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Western blot experiments were performed exactly as described [29 (link)]. Blots were probed mouse anti-FAAH1 (1:1000, Abcam #Ab54615), mouse anti-GAPDH (1:5000, Abcam #Ab8245), mouse anti-FLAG (1:1000, Sigma #F1804), or rabbit anti-FAAH2 (1:200, Abcam #Ab103724) antibodies. The blots were then incubated with the respective goat anti-mouse or goat anti-rabbit IgG HRP-conjugated antibodies (Life Technologies), developed using the Immun-star HRP substrate (Bio-Rad), and scanned using a C-DiGiT scanner (Li-COR).
Sirrs S., van Karnebeek C.D., Peng X., Shyr C., Tarailo-Graovac M., Mandal R., Testa D., Dubin D., Carbonetti G., Glynn S.E., Sayson B., Robinson W.P., Han B., Wishart D., Ross C.J., Wasserman W.W., Hurwitz T.A., Sinclair G, & Kaczocha M. (2015). Defects in fatty acid amide hydrolase 2 in a male with neurologic and psychiatric symptoms. Orphanet Journal of Rare Diseases, 10, 38.
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5

Western Blot Analysis of Alzheimer's Biomarkers

Cited 2 times
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Brain homogenates (frontal cortex) in urea (50 μg) were mixed with Laemmli sample buffer and resolved by SDS-PAGE before an overnight wet transfer to nitrocellulose membranes (BioRad) as previously reported (Seyfried et al., 2017 (link)). Membranes were blocked with casein blocking buffer (Sigma B6429) and probed with primary polyclonal antibodies for Anti-PLCD1 antibody (Abcam #ab134936), anti-HEPACAM (Abcam #ab130769), anti-PADI2 antibody (Abcam #ab16478), anti-GAPDH antibody (Abcam #ab8245), and anti-Connexin-43 (Sigma #C6219) overnight at 4°C. Membranes were incubated with secondary antibodies conjugated to Alexa Fluor 680 (Invitrogen) or IRDye800 (Rockland) fluorophores for one hour at room temperature. Images were captured using an Odyssey Infrared Imaging System (Li-Cor Biosciences). Protein densitometry for relative quantification was performed using ImageJ open source software. For GJA1, the higher molecular weight band reflective of ubiquitination (Abreha et al., 2018 (link)) was analyzed separately from the lower molecular weight band. Densitometry was compared between AD and control using the Student’s t test (Student, 1908 ).
Swarup V., Chang T.S., Duong D.M., Dammer E.B., Dai J., Lah J.J., Johnson E.C., Seyfried N.T., Levey A.I, & Geschwind D.H. (2020). Identification of Conserved Proteomic Networks in Neurodegenerative Dementia. Cell reports, 31(12), 107807.
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6

Antibody Usage in Cell Biology Research

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Antibodies used were as follows: against α-tubulin, from Abcam (Cambridge, UK), ab7291 and ab18251, made in mouse and rabbits, respectively (used at dilutions of 1:2000 for Western blot and 1:200 for microscopy); against Dia 1 (1:1000), ab1173; against glyceraldehyde phosphate dehydrogenase, ab8245 (1:40,000); against kinesin, K1005 (1:50); against Ac-tubulin, T6793 (1:500); against the profilin I N-terminal, P7749 (1:100); and against β-actin, A5441 (1:4000), from Sigma-Aldrich, Stockholm, Sweden; against profilin I (our laboratory, raised in rabbits, 1:2000); against β-arrestin, from Santa Cruz Biotechnology (Heidelberg, Germany), against Dia 2, sc9182 (1:20) and sc-393499 (1:100); against GFP, from Roche (Stockholm, Sweden), 11814460001 (1:2000); horseradish peroxidase (HRP) conjugated against mouse immunoglobulin (Ig), from Dako (Stockholm, Sweden), P0447 (1:2000); against rabbit Ig, from Pierce (Stockholm, Sweden), 1858415 (1:1000); fluorescein isothiocyanate (FITC) conjugated against mouse and rabbit Ig, from Jackson ImmunoResearch Laboratories (West Grove, PA), 705-095-003 (1:400) and 711-095-152 (1:1000), respectively; and tetramethylrhodamine isothiocyanate conjugated against rabbit Ig, from Thermo Fisher Scientific (Stockholm, Sweden), 115-025-062 (1:400).
Nejedla M., Sadi S., Sulimenko V., de Almeida F.N., Blom H., Draber P., Aspenström P, & Karlsson R. (2016). Profilin connects actin assembly with microtubule dynamics. Molecular Biology of the Cell, 27(15), 2381-2393.
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7

Comprehensive Protein Expression Profiling

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ES cells were lysed in RIPA buffer supplemented with protease inhibitors (11836170001; Roche). Twelve percent SDS-PAGE gels were used for protein separation followed by transfer onto PVDF membrane. Blots were blocked with 5% non-fat milk at room temperature for 1 h and incubated with the following primary antibodies overnight at 4 °C: RNF126 (Abcam, ab234812, 1:500), BHMT (Abcam, ab96415, 1:500), NQO1 (Abcam, ab28947, 1:500), KRAS (Abcam, Ab180772, 1:500), S100A6 (Abcam, ab134149, 1:1000), OCT4 (Millipore, MABD76, 1:1000), and ESRRB (Perseus proteomics, PP-H6705-00, 1:1000). Primary antibodies for GAPDH (Abcam, Ab8245, 1:1000) and ACTB (Sigma, A1978, 1:1000) were employed as internal controls and for 1 h at room temperature. HRP-swine-anti-rabbit (Dako, P0217) and HRP-rabbit anti mouse (Dako, P0161) were used as secondary antibodies and subsequently signal was detected using ECL kit (Pierce, 32106) and the ImageQuant LAS 4000 system (GE Healthcare Life Sciences). Western blots were repeated for two times.
Atlasi Y., Jafarnejad S.M., Gkogkas C.G., Vermeulen M., Sonenberg N, & Stunnenberg H.G. (2020). The translational landscape of ground state pluripotency. Nature Communications, 11, 1617.
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8

Immunoblotting of PKM Silenced Cells

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Cells transfected with Pkm siRNA were harvested 4 d after transfection. Preparation of whole-cell extracts and immunoblotting were done as described.25 (link) Antibodies used were against Ser473 phosphorylated AKT (Cell Signaling Technologies, 4058), GAPDH (Abcam, Ab8245) and PKM (Sigma-Aldrich, SAB4200095).
Isidor M.S., Winther S., Basse A.L., Petersen M.C., Cannon B., Nedergaard J, & Hansen J.B. (2015). An siRNA-based method for efficient silencing of gene expression in mature brown adipocytes. Adipocyte, 5(2), 175-185.
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9

Immunodetection of Brain Proteins

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Antibodies against the following proteins were used in Western blots and for immunofluorescence analysis: calbindin (mouse, Swant, 300, dilution 1:500), CB1 (rabbit, Frontier Institute, AF380, 1:500), CB2 (mouse, R&D Systems, 352110, 1:500), F4/80 (rat, Abcam, ab6640, 1:500), GAPDH (mouse, Abcam, ab8245, 1:5,000), GFAP (mouse, Millipore, MAB3402, 1:1,000), Iba1 (rabbit, Wako, 019‐19741, 1:500), Lamp1 (rat, DSHB, 1D4B, 1:500), MAP2 (chicken, BioLegend, 822501, 1:500), NSM (rat, Santa Cruz, sc‐166637, 1:200), PSD‐95 (mouse, BD Transduction Laboratories, 610495, 1:500), and cleaved caspase 3 (Asp 175; rabbit polyclonal, Cell Signaling, 9661, 1:200). HRP‐conjugated rabbit anti‐mouse or anti‐rat and goat anti‐rabbit (DakoCytomation) were used as secondary antibodies in Western blots. Alexa‐conjugated rabbit anti‐mouse or anti‐rat and goat anti‐rabbit were used as secondary antibodies in immunofluorescence.
Bartoll A., Toledano‐Zaragoza A., Casas J., Guzmán M., Schuchman E.H, & Ledesma M.D. (2020). Inhibition of fatty acid amide hydrolase prevents pathology in neurovisceral acid sphingomyelinase deficiency by rescuing defective endocannabinoid signaling. EMBO Molecular Medicine, 12(11), e11776.
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10

Western Blot Analysis of Chromatin Regulators

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Cells were lysed in RIPA buffer and lysates were separated by SDS-PAGE and transferred to PVDF paper. Antibodies used were FMRP (Cell signaling 4317S, 1:1000), Brd4 (Bethyl A301-985A, 1:1000), phospho-Brd4 (developed with Millipore, described in (Korb et al., 2015 (link)), 1:500), p300 (sc-585, 1:300), MLL1 (Bethyl A300-374, 1:500), Gapdh (Abcam ab8245, 1:500), GluA1 (Millipore MAB2263, 1:500), Shank2 (Cell Signaling 12218, 1:500), BDNF (Abcam ab88901, 1:500), CaMK2alpha (Millipore 05-532, 1:500), H3 (Abcam ab1791, 1:4000), H4 (Abcam ab10158, 1:4000), H3K4me3 (Active motif 39159, 1:3000), H4K8ac (Millipore 07-328, 1:1000), H4K16ac (Active motif 39167, 1:500), H3K27ac (Active motif 39133, 1:1000), and H3K27me3 (abcam ab6002, 1:500). Western blots were quantified normalizing each lane by the Gapdh signal to control for loading differences.
Korb E., Herre M., Zucker-Scharff I., Gresack J., Allis C.D, & Darnell R.B. (2017). Excess translation of epigenetic regulators contributes to Fragile X Syndrome and is alleviated by Brd4 inhibition. Cell, 170(6), 1209-1223.e20.
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