Anti mouse ig h l

Manufactured by Southern Biotech

Anti-mouse Ig(H+L) is a laboratory reagent used to detect and quantify mouse immunoglobulins (Ig) in various immunoassays. It binds to the heavy and light chains of mouse Ig molecules, allowing for their identification and measurement.

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Lab products found in correlation

Anti mouse igm ap, by Southern Biotech (2 mentions) Anti mouse igg1 ap, by Southern Biotech (2 mentions) Np25 bsa, by Southern Biotech (2 mentions) Np5 bsa, by Southern Biotech (2 mentions) Alkaline phosphatase conjugated anti igm or anti igg1 antibodies, by Southern Biotech (2 mentions) Complete freund s adjuvant, by Merck Group (2 mentions) Goat anti mouse ig h l, by Southern Biotech (2 mentions) Bcip nbt, by Merck Group (2 mentions) Streptavidin alkaline phosphatase, by Merck Group (2 mentions) Streptavidin conjugated horseradish peroxidase sa hrp and tmb substrate, by BD (1 mentions) Np25 bsa, by Biosearch Technologies (1 mentions) Np9 bsa, by Biosearch Technologies (1 mentions) P nitrophenylphosphate, by Southern Biotech (1 mentions) Nbt bcip, by Roche (1 mentions) 96 well immune plate, by Thermo Fisher Scientific (1 mentions) Opteia tmb substrate, by BD (1 mentions) Msips4w10, by Merck Group (1 mentions) L α phosphatidylcholine, by Merck Group (1 mentions) Bovine serum albumin (bsa), by Nacalai Tesque (1 mentions) Tmb peroxidase substrate, by LGC (1 mentions)

Spelling variants of this product

Goat anti-mouse Ig (H+L) (17 citations)

9 protocols using anti mouse ig h l

Rapid ELISA for WNV Antibodies

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Detection of WNV-specific antibodies was based on ELISAs with recombinant WNV-E, DIII, or DIII-KT as antigens. Production of these reagents followed published protocols [77 (link),78 (link)]. Briefly, DIII proteins were expressed in BL21(DE3) E. coli cells, refolded from inclusion bodies by oxidative refolding, and purified by size exclusion. AviTag-DIII protein was biotinylated and purified again by size exclusion. Serial dilutions of serum from RWN-infected mice were applied to plates coated with recombinant WNV-E, DIII; or DIII-KT proteins. Bound RWN-specific antibodies were detected with biotinylated goat anti-mouse IgM, IgG, IgG2b, IgG2c, or IgG3 (Southern Biotech, Birmingham, AL), followed by streptavidin-conjugated horseradish peroxidase (SA-HRP) and TMB substrate (both BD Bioscience, San Diego, CA). Anti-mouse Ig(H+L) (Southern Biotech, Birmingham, AL) and serial dilutions of mouse IgM and IgG2c (Southern Biotech, Birmingham, AL) were used for standards. High-affinity antibodies were measured similarly using plates coated with recombinant DIII alone or diluted 1:3 with BSA. After the initial binding, low affinity antibodies were washed off by incubating the samples for 15 min in presence of increasing amounts of NaSCN before detection with biotinylated goat anti-mouse IgM or IgG, followed by SA-HRP.

O’Ketch M., Williams S., Larson C., Uhrlaub J.L., Wong R., Hall B., Deshpande N.R, & Schenten D. (2020). MAVS regulates the quality of the antibody response to West-Nile Virus. PLoS Pathogens, 16(10), e1009009.

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ELISA and ELISPOT Assays for Antibody Responses

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For SRBC immunization experiments, 96-well immune plates (Thermo Fisher Scientific) were coated with anti–mouse Ig(H+L) (Southern Biotech). For NP immunization experiments, plates were coated with NP₉-BSA or NP₂₅-BSA (Biosearch Technologies). Mouse serum samples were incubated for 2 h at room temperature. Standard curves were generated using mouse IgM and mouse IgG1 (Southern Biotech). Bound antibodies were detected by AP-conjugated anti–mouse IgM and anti–mouse IgG1 antibodies (Table S5). Plates were developed with p-nitrophenylphosphate (Southern Biotech) dissolved in substrate buffer. For ELISPOT analysis, the spleens and BM of NP-KLH–immunized mice were removed at the indicated time points, and single cell suspensions were subjected to hypotonic lysis. The samples were plated overnight on 96-well filtration plates (Millipore) coated with NP₂₅-BSA (Biosearch Technologies). The cells were plated in 1:2 serial dilutions, starting at 8 × 10⁵ cells for BM-derived samples, and at 10⁵ cells for spleen-derived fractions. NP-specific IgG1-secreting cells were detected by AP-conjugated anti–mouse IgG1 antibody (Table S5). Plates were developed with nitro blue tetrazolium chloride-5-bromo-4-chloro-3-indolyl phosphate (NBT/BCIP; Roche).

Heise N., De Silva N.S., Silva K., Carette A., Simonetti G., Pasparakis M, & Klein U. (2014). Germinal center B cell maintenance and differentiation are controlled by distinct NF-κB transcription factor subunits. The Journal of Experimental Medicine, 211(10), 2103-2118.

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NP-Specific Antibody Titer Quantification

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Total IgM and IgG1 serum titers were measured by ELISA as previously described^{65 (link)}, using the following antibodies: anti-Mouse Ig H+L (1010-01, Southern Biotech); anti-Mouse IgM-AP (1020-04, Southern Biotech); and anti-Mouse IgG1-AP (1070-04, Southern Biotech). To quantify NP-specific antibodies, serum was collected at day 0, 12 and 28 after a single intraperitoneal injection of NP-KLH (100 μg) in complete Freund's adjuvant (Sigma). NP-specific high affinity and low affinity antibodies were captured on plates coated with 2 μg/ml NP₂₅-BSA or NP₅-BSA, respectively (SouthernBiotech), and bound antibodies were detected with alkaline phosphatase-conjugated anti-IgM or anti-IgG1 antibodies (SouthernBiotech). Pan-mouse immunoglobulins were captured on plates coated with 2.5 μg/ml goat anti-mouse Ig (H+L) (SouthernBiotech). Each assay was performed in duplicate using twelve serial dilutions.

Zhang J., Dominguez-Sola D., Hussein S., Lee J.E., Holmes A.B., Bansal M., Vlasevska S., Mo T., Tang H., Basso K., Ge K., Dalla-Favera R, & Pasqualucci L. (2015). Disruption of KMT2D perturbs germinal center B cell development and promotes lymphomagenesis. Nature medicine, 21(10), 1190-1198.

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Quantifying Antibody Responses via ELISA and ELISpot

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ELISA and ELISpot assays were performed according to standard protocols. Briefly, plates (ThermoFisher 442404) were coated with anti-mouse Ig(H+L) (Southern Biotech 1010-01) in a sodium carbonate/bicarbonate solution pH 9.6. Helicobacter ELISpot plates were coated with 200ng/well of heat-killed H. muridarum. Serum was serially diluted and detected using an HRP conjugated anti-mouse IgA antibody (Sigma A4789) and BD OptEIA TMB substrate (BD 555214). ELISpot plates (EMD Millipore MSIPS4W10) were coated the same as ELISA plates, but detection was performed with anti- IgM, IgA, IgG, κ, and λ antibodies conjugated to biotin (Southern Biotech). biotinylated antibodies were revealed with streptavidin-alkaline phosphatase (Sigma E2636) and developed with BCIP/NBT (Sigma B1911). ELISpot plates were imaged and spots were counted using a CTL Immunospot Analyzer (Cellular Technologies Limited).

Wilmore J.R., Gaudette B.T., Atria D.G., Hashemi T., Jones D.D., Gardner C.A., Cole S.D., Misic A.M., Beiting D.P, & Allman D. (2018). Commensal microbes induce serum IgA responses that protect against polymicrobial sepsis. Cell host & microbe, 23(3), 302-311.e3.

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Quantification of Antibody and Cytokine Responses

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ELISAs were performed as previously described (48 (link)). Samples were diluted in TBS containing 1% BSA (TBS-BSA) and analyzed for total and PC-specific IgM, IgG, and IgA. Diluted serum samples were added to Costar plates coated with 0.625 μg/mL PC(4 (link))BSA (Biosearch Technologies) or 5 μg/mL anti-mouse Ig(H+L) (Southern Biotechnology Associates) or AB1.2 (anti-T15 Id) Ab and blocked with TBS-BSA. Alkaline phosphatase-conjugated polyclonal goat anti-mouse IgM, IgG, IgG3, and IgA Abs (Southern Biotech) and pNPP (Sigma) were used to detect Ab. A PC-specific IgM hybridoma was used for quantitation of the PC IgM response. IL-4, IL-5, IL-6, IL-10, IFNγ, TNFα, and GM-CSF ELISAs were performed according to manufacturer’s instructions (BioLegend). Malondialdehyde-specific ELISAs used plates coated with MDA-BSA (Cell Biolabs, Inc) at 5 ug/ml. Phosphatidylcholine ELISAs were performed on air-dried plates that had been coated with L-alpha-phosphatidylcholine (Sigma) at 30 ug/ml in hexane. ELISPOT was performed as previously described (33 (link)), using both Image Acquisition 4.5 and ImmunoSpot software for analysis of spot number and size.

McKay J.T., Haro M.A., Daly C.A., Yammani R.D., Pang B., Swords W.E, & Haas K.M. (2017). PD-L2 regulates B-1 cell antibody production against phosphorylcholine through an IL-5–dependent mechanism. Journal of immunology (Baltimore, Md. : 1950), 199(6), 2020-2029.

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Quantification of Antibody Responses to E. coli

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As previously described, E. coli strain S-17^{76 (link)} were freeze-dried by Techno Suruga Laboratory. Ninety-six-well plates were coated with the anti-mouse Ig(H + L) (1 μg/mL; Southern Biotech, #1010-01) to detect total IgA or freeze-dried S-17 (1 mg/mL) for detection of anti-E. coli IgA, IgM, IgG and incubated at 4 °C overnight. The plates were washed, 1% bovine serum albumin (Nacalai Tesque) in PBS was added for blocking, and the plates were incubated for 1 h at room temperature. After the plates were washed, test samples were added, and the plates were incubated overnight at 4 °C. Then, the plates were washed and horseradish peroxidase-conjugated anti-mouse IgA, IgM, IgG (Southern Biotech, #1040-05, #1040-05, #1020-05, 1:4000) in 1% bovine serum albumin in PBS was added, respectively, and the plates were incubated for a further 1 h at room temperature. The plates were then washed, and TMB Peroxidase Substrate (Kirkegaard & Perry Laboratories, #5120-0050) was added. Stop solution was added after 2 min incubation, followed by reading at 450 nm with a spectrometer. A GP2 ELISA kit (MyBioSource, #MBS081334) was used to detect fecal GP2, in accordance with the manufacturer’s instructions.

Kurashima Y., Kigoshi T., Murasaki S., Arai F., Shimada K., Seki N., Kim Y.G., Hase K., Ohno H., Kawano K., Ashida H., Suzuki T., Morimoto M., Saito Y., Sasou A., Goda Y., Yuki Y., Inagaki Y., Iijima H., Suda W., Hattori M, & Kiyono H. (2021). Pancreatic glycoprotein 2 is a first line of defense for mucosal protection in intestinal inflammation. Nature Communications, 12, 1067.

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Quantitative IgG and IL-22 ELISA Protocols

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For IgG ELISAs, a 96-well high-binding assay plate (Corning) was coated with either 10 µg/ml of unlabeled polyclonal anti-mouse Ig (H⁺L) for total IgG ELISA or 5 µg/ml of recombinant Intimin protein for C.r-specific IgG ELISA in 1x PBS overnight at 4°C. After 4 washes with 1x PBS, 1% BSA in 1x PBS Blocking solution was added to the plate and incubated at RT for 1 hr. After 4 washes with 1x PBS/Tween-20, samples were diluted in 1% BSA/PBS/Tween-20 and incubated at RT for 2 hrs. After 4 washes with 1x PBS/Tween-20, 1:4000 dilution of anti-mouse IgG (H⁺L)-HRP conjugated was incubated at RT for 2 hrs. After the final 4 washes, 100 µl of a TMB single solution (Life Tech) was added to the plate and the chemiluminescence signal was stopped with 2N Sulfuric acid and read at 450 nm. All samples were run in triplicate and IgG standards were included on all plates. The following antibodies purchased from Southern Biotech were used: anti-mouse Ig (H⁺L) unconjugated (for coating plate), purified mouse IgG (for standard) and anti-mouse IgG HRP labeled (for detection). For IL-22 protein quantitation, a mouse/rat IL-22 Quantikine ELISA kit (R&D Systems, M2200) was used according to manufacturer’s protocol. See details in Key Resources Table.

Zindl C.L., Witte S.J., Laufer V.A., Gao M., Yue Z., Janowski K.M., Cai B., Frey B.F., Silberger D.J., Harbour S.N., Singer J.R., Turner H., Lund F.E., Vallance B.A., Rosenberg A.F., Schoeb T.R., Chen J.Y., Hatton R.D, & Weaver C.T. (2022). A Non-redundant Role for T cell-derived Interleukin 22 in Antibacterial Defense of Colonic Crypts. Immunity, 55(3), 494-511.e11.

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NP-Specific Antibody Titer Quantification

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ELISpot Assay for Antibody Detection

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ELISpot assays were performed according to standard protocols. Briefly, ELISpot plates (EMD Millipore MSIPS4W10) were coated with anti-mouse Ig(H+L) (Southern Biotech 1010-01) in a sodium carbonate/bicarbonate solution pH 9.6 and detection was performed with anti- IgM, IgA, IgG, κ, and λ antibodies conjugated to biotin (Southern Biotech). Biotinylated antibodies were revealed with streptavidin-alkaline phosphatase (Sigma E2636) and developed with BCIP/NBT (Sigma B1911). ELISpot plates were imaged and spots were counted using a CTL Immunospot Analyzer (Cellular Technologies Limited).

Wilmore J.R., Gaudette B.T., Gómez Atria D., Rosenthal R.L., Reiser S.K., Meng W., Rosenfeld A.M., Luning Prak E.T, & Allman D. (2021). IgA Plasma Cells Are Long-Lived Residents of Gut and Bone Marrow That Express Isotype- and Tissue-Specific Gene Expression Patterns. Frontiers in Immunology, 12, 791095.

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