Anti human igg apc

Manufactured by BD

Anti-human IgG-APC is a fluorescently labeled antibody that binds to human immunoglobulin G (IgG) antibodies. It is used in flow cytometry and other immunoassays to detect and quantify the presence of human IgG in a sample.

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Lab products found in correlation

Rnaseout, by Thermo Fisher Scientific (3 mentions) Dithiothreitol (dtt), by Thermo Fisher Scientific (3 mentions) Rnasin, by Promega (3 mentions) Anti human cd27 pe, by BD (3 mentions) Facsaria fusion, by BD (3 mentions) Anti human cd20 alexa fluor 700, by BD (2 mentions) Cd19 microbeads, by Miltenyi Biotec (2 mentions) Isotype control apc, by BD (2 mentions) 4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (2 mentions) Pan b cell isolation kit, by Miltenyi Biotec (1 mentions) B cell isolation kit 2, by Miltenyi Biotec (1 mentions) Igg memory b cell isolation kit, by Miltenyi Biotec (1 mentions) Streptavidin pe, by BD (1 mentions) 4 6 diamidin 2 phenylindol dapi, by Thermo Fisher Scientific (1 mentions) Cytofix buffer, by BD (1 mentions) Lsrfortessa flow cytometer, by BD (1 mentions) Anti human cd19 pecy7, by BD (1 mentions) Ficoll density gradient centrifugation, by GE Healthcare (1 mentions) Facsvantage cell sorter, by BD (1 mentions)

6 protocols using anti human igg apc

Single-cell Sorting of HIV-1 Env-Reactive B Cells

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B cells were isolated from PBMCs using the Pan B Cell Isolation Kit, B Cell Isolation Kit II, or IgG⁺ Memory B Cell Isolation Kit (Miltenyi Biotec). Isolated cells were labeled with anti-human CD19-AF700 (BD), anti-human IgG-APC (BD), DAPI (Thermo Fisher), and the respective HIV-1 Env bait for 30 minutes on ice. BG505_SOSIP.664-GFP or biotinylated YU2_gp140 that was labeled with Streptavidin-PE (BD) were used as HIV-1 Env baits. Env-reactive CD19⁺IgG⁺DAPI^- single cells were sorted into 96-well plates containing 4 μl of lysis buffer (0.5x PBS, 10 mM DTT (Thermo Fisher), 2 U/μl RNasin (Promega), and 1 U/μl RNaseOUT (Thermo Fisher)) per well using a BD FACSAria Fusion. Plates were stored at −80°C until further use.

Schommers P., Gruell H., Abernathy M.E., Tran M.K., Dingens A.S., Gristick H.B., Barnes C.O., Schoofs T., Schlotz M., Vanshylla K., Kreer C., Weiland D., Holtick U., Scheid C., Valter M.M., van Gils M.J., Sanders R.W., Vehreschild J.J., Cornely O.A., Lehmann C., Fätkenheuer G., Seaman M.S., Bloom J.D., Bjorkman P.J, & Klein F. (2020). Restriction of HIV-1 Escape by a Highly Broad and Potent Neutralizing Antibody. Cell, 180(3), 471-489.e22.

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Isolation and Characterization of SARS-CoV-2-Specific B Cells

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B cells were isolated from PBMCs using CD19-microbeads (Miltenyi Biotec) according to the manufacturer’s instruction. Isolated B cells were stained for 20 min on ice with a fluorescence staining-mix containing 4’,6-Diamidin-2-phenylindol (DAPI; Thermo Fisher Scientific), anti-human CD20-Alexa Fluor 700 (BD), anti-human IgG-APC (BD), anti-human CD27-PE (BD) and DyLight488-labeled SARS-CoV-2 spike protein (10μg/mL). Dapi⁻, CD20⁺, IgG⁺, SARC-CoV-2 spike protein positive cells were sorted using a FACSAria Fusion (Becton Dickinson) in a single cell manner into 96-well plates. All wells contained 4 μL buffer, consisting of 0.5x PBS, 0.5 U/μL RNAsin (Promega), 0.5 U/μL RNaseOUT (Thermo Fisher Scientific), and 10 mM DTT (Thermo Fisher Scientific). After sorting, plates were immediately stored at −80°C until further processing.

Kreer C., Zehner M., Weber T., Ercanoglu M.S., Gieselmann L., Rohde C., Halwe S., Korenkov M., Schommers P., Vanshylla K., Di Cristanziano V., Janicki H., Brinker R., Ashurov A., Krähling V., Kupke A., Cohen-Dvashi H., Koch M., Eckert J.M., Lederer S., Pfeifer N., Wolf T., Vehreschild M.J., Wendtner C., Diskin R., Gruell H., Becker S, & Klein F. (2020). Longitudinal Isolation of Potent Near-Germline SARS-CoV-2-Neutralizing Antibodies from COVID-19 Patients. Cell, 182(4), 843-854.e12.

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BHK21 Cell Immunophenotyping Protocol

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Forty-two hours after transfection, BHK21 cells were harvested, washed with two to four volumes of staining buffer (1% BSA in PBS), and centrifuged (500 ×g, 5 min, 4°C). Five hundred thousand cells were resuspended in 80 μl of staining buffer and 20 μl of diluted antibodies added (5 μg per mL, 1 hour, 4°C). Cells were washed and incubated in 80 μl of staining buffer with 20 μl isotype-APC control (BD cat #555751) or 20 μl anti-Human IgG-APC (BD cat #550931) for 30–45 minutes at 4°C. Cells were washed and fixed with 250 μl of BD Cytofix Buffer (Cat #554655) for 30 minutes at 4°C, and were analyzed on a BD LSRFortessa flow cytometer.

Yu J.S., Liao H.X., Pritchett J., Bowman C., Vivian C., Parks R., Xia S.M., Cooper M., Williams W.B., Bonsignori M., Reed S.G., Chen M., Vandergrift N., Rice C.M, & Haynes B.F. (2017). Development of a Recombinant Yellow Fever Vector Expressing a HIV Clade C Founder Envelope gp120. Journal of virological methods, 249, 85-93.

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SARS-CoV-2 Spike Protein B Cell Sorting

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CD19⁺ B cells were enriched from PBMCs by immunomagnetic cell separation using CD19 microbeads (Miltenyi Biotec) according to the manufacturer’s protocol. Enriched B cells were labeled for 20 min on ice with 4’,6-Diamidin-2-phenlindol (DAPI; Thermo Fisher Scientific), anti-human CD20-Alexa Fluor 700 (BD), anti-human IgG-APC (BD), anti-human CD27-PE (BD) and DyLight488-labeled SARS-CoV-2 S protein (10 μg/ml). DAPI⁻, CD20⁺, SARS-CoV S protein⁺, IgG⁺/IgG⁻ single cells were sorted into 96 well plates using a FACSAria Fusion (Becton Dickinson). Each well of the 96 well plate was prefilled with 4 μl of sorting buffer consisting of 0.5 x PBS, 0.5 U/μl RNAsin (Promega), 0.5 U/μl RNaseOut (Thermo Fisher Scientific), and 10 mM DTT (Thermo Fisher Scientific). Plates were cryopreserved at −80°C immediately after sorting.

Ercanoglu M.S., Gieselmann L., Dähling S., Poopalasingam N., Detmer S., Koch M., Korenkov M., Halwe S., Klüver M., Di Cristanziano V., Janicki H., Schlotz M., Worczinski J., Gathof B., Gruell H., Zehner M., Becker S., Vanshylla K., Kreer C, & Klein F. (2022). No substantial preexisting B cell immunity against SARS-CoV-2 in healthy adults. iScience, 25(3), 103951.

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Surface Expression of VRC-A gp145 in 293i Cells

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For the surface expression of VRC-A gp145, 293i cells were transfected
with pHV130770 plasmids using ExpiFectamine™ 293 Transfection Kits (Life
Technologies Cat. # A14525) following the manufactures’
protocol. The cells were washed with four volumes of room-temperature
phosphate-buffered saline (DPBS) (Life Technologies Cat # 10010-023),
and centrifuged (500g for 5 minutes). 10⁶ cells were resuspended in
80 µL of staining buffer (1% BSA in DPBS) to which were added 20
µL of diluted antibodies (1 ug per reaction). After incubation (1 hour
at 4°C) with shaking, cells were washed and centrifuged (500 X g). Cells
were incubated with either 20 µL isotype-APC control (BD cat
#555751) or 20 µL anti-Human IgG-APC (BD cat #550931)
for 30–45 minutes at 4°C, protected from light. After washing
and fixation, cells were analyzed by flow cytometer (BD LSRFortessa).

Williams W.B., Liao H.X., Moody M.A., Kepler T.B., Alam S.M., Gao F., Wiehe K., Trama A.M., Jones K., Zhang R., Song H., Marshall D.J., Whitesides J.F., Sawatzki K., Hua A., Liu P., Tay M.Z., Seaton K., Shen X., Foulger A., Lloyd K.E., Parks R., Pollara J., Ferrari G., Yu J.S., Vandergrift N., Montefiori D.C., Sobieszczyk M.E., Hammer S., Karuna S., Gilbert P., Grove D., Grunenberg N., McElrath J., Mascola J.R., Koup R.A., Corey L., Nabel G.J., Morgan C., Churchyard G., Maenza J., Keefer M., Graham B.S., Baden L.R., Tomaras G.D, & Haynes B.F. (2015). Diversion of HIV-1 Vaccine-induced Immunity by gp41-Microbiota Cross-reactive Antibodies. Science (New York, N.Y.), 349(6249), aab1253.

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Intestinal plasmablast/plasma cell isolation

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Intestinal plasmablasts/plasma cells were isolated as previously described (Benckert et al., 2011 (link)). In brief, lamina mucosa and propria were dissected from lamina muscularis mucosae by blunt preparation, and 2–3-mm tissue pieces were digested using 0.1% DNase and 0.1% collagenase followed by discontinuous Ficoll density gradient centrifugation (GE Healthcare). Purified lamina propria lymphocytes were stained on ice with fluorochrome-coupled anti-human CD38 FITC, anti-human CD27 PE, anti-human CD19 PECy7, anti-human IgG APC (all from BD Bioscience) or anti-human IgA APC (Jackson Laboratory). Single CD38⁺CD27⁺IgA⁺ or CD38⁺CD27⁺IgG⁺ plasmablasts/plasma cells were sorted into 96-well PCR plates using a FACSVantage cell sorter with Diva configuration (BD Bioscience), snap frozen on dry ice, and stored at −80°C until further processing.

Kabbert J., Benckert J., Rollenske T., Hitch T.C., Clavel T., Cerovic V., Wardemann H, & Pabst O. (2020). High microbiota reactivity of adult human intestinal IgA requires somatic mutations. The Journal of Experimental Medicine, 217(11), e20200275.

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