Purified mouse igg1

Manufactured by BD

Sourced in United States

Purified mouse IgG1 is a laboratory reagent that provides a source of immunoglobulin G1 (IgG1) derived from mice. IgG1 is a class of antibodies that can be used in various immunological applications.

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4 protocols using purified mouse igg1

Quantification of Mouse Serum Immunoglobulins

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The OptEIA Mouse IgE set (BD Pharmingen^TM, Erembodegem, Belgium) was used to measure total serum IgE (diluted 1/70). Measurements were performed according to the manufacturer's instructions. For the measurement of total serum IgG1 (diluted 1/8,000 or 1/20,000) and IgG2a (diluted 1/1,000), plates were coated using purified rat anti-mouse IgG1 (Cat: 553445; BD Pharmingen^TM) and rat anti-mouse IgG2a (Cat: 553446; BD Pharmingen^TM). A standard was created using purified mouse IgG1 (Cat: 557273; BD Pharmingen^TM) and mouse IgG2a (Cat: 553454; BD Pharmingen^TM). Further measurements were performed according to the manufacturer's instructions with the use of biotinylated anti-mouse avidin horseradish peroxidase (HRP) conjugate (HRP rat anti-mouse IgG1 [Cat: 559626], biotin rat anti-mouse IgG2a [Cat: 553388] and Streptavidin HRP [Cat: 554066]; BD Pharmingen^TM).

Pollaris L., Decaesteker T., Van den Broucke S., Jonckheere A.C., Cremer J., Verbeken E., Maes T., Devos F.C., Vande Velde G., Nemery B., Hoet P.H, & Vanoirbeek J.A. (2020). Involvement of Innate Lymphoid Cells and Dendritic Cells in a Mouse Model of Chemical-induced Asthma. Allergy, Asthma & Immunology Research, 13(2), 295-311.

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Isolation and Characterization of Human Islet Cells

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Isolated human islets were digested with Accutase (Sigma-Aldrich) to generate a single cell population, which were then stained with a viability dye 7-AAD (Life Technologies), an isotype control (purified mouse IgG1, BD Biosciences, New Jersey, USA), mouse anti-human DSG2 (clone 6D8, Life Technologies) for 30 min at 4 °C. Cells were washed twice then stained with a secondary antibody; goat anti-mouse-DyLight 650 (Abcam) as well as a β-cell marker Newport Green (Invitrogen; Thermo Fisher Scientific) for 30 min at 4 °C. Cells were washed and then resuspended in FACS fix (1% formaldehyde, 20 g/L glucose, 5 mM sodium azide in PBS) and analyzed on the BD Accuri C6 flow cytometer (BD Biosciences). Data was further analyzed using FCS Express 4 Flow Cytometry: Research Edition (De Novo Software).

Myo Min K.K., Rojas-Canales D., Penko D., DeNichilo M., Cockshell M.P., Ffrench C.B., Thompson E.J., Asplund O., Drogemuller C.J., Prasad R.B., Groop L., Grey S.T., Thomas H.E., Loudovaris T., Kay T.W., Mahoney M.G., Jessup C.F., Coates P.T, & Bonder C.S. (2022). Desmoglein-2 is important for islet function and β-cell survival. Cell Death & Disease, 13(10), 911.

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Quantitative ELISA for Mouse Immunoglobulins

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Immulon 4 plates (Thermo Scientific, Waltham, MA) were coated with either purified mouse anti-IgE capture antibody (BD Biosciences), purified mouse anti-IgG1 (BD Pharmingen, Franklin Lakes, NJ) capture antibody or purified mouse anti-IgG2 (BD Pharmingen) capture antibody in 1X PBS and incubated overnight at 4°C. Then plates were washed with 1X PBS supplemented with 0.025% Tween 20 and blocked for 2 hours at 37 °C in 1X PBS supplemented with 5% BSA (Sigma Aldrich) and 0.05% Tween. Purified mouse IgE (BD Biosciences), purified mouse IgG1 (BD Biosciences) and purified mouse IgG2 (BD Biosciences) were used as standards. Blocked plates were washed with wash buffer and samples incubated for 2 hours at 37 °C. Plates were then washed and biotinylated anti-mouse IgE (BD Biosciences), anti-mouse IgG1 (BD Biosciences) or anti-mouse IgG2 (BD Biosciences) was added and incubated for 2 hours at 37 °C. Following a washing step, 1:1000 dilution of streptavidin conjugated alkaline phosphatase (Jackson Immunoresearch, West Grove, PA) was added to the plates and incubated for 1 hour at 37 °C. 4-Nitrophenyl phosphate disodium salt hexahydrate (PNPP) (Sigma Aldrich) was used as the substrate. Plates were read on an ELISA reader at 405/650.

Boyd A., Killoran K., Mitre E, & Nutman T.B. (2015). Pleural cavity Type 2 innate lymphoid cells precede Th2 expansion in murine Litomosoides sigmodontis infection. Experimental parasitology, 159, 118-126.

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Quantifying Murine Antibody Responses

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For serum OVA-specific IgG1 antibody detection, Nunc Maxisorp 96-well plates (Thermo Fisher Scientific) were coated with 10 μg/ml OVA (Grade V, Sigma-Aldrich) O/N at 4°C, subsequently blocked with 1 % BSA and serial dilutions of sera were added (1:1000, 1:2500, 1:5000, 1:10000). Purified mouse IgG1 (BD Biosciences) was used as standard (starting concentration 500 ng/ml). Detection was achieved with biotinylated anti-IgG1 (BD Biosciences) at 0.1 μg/ml. For total IgE detection, the plates were coated with anti-mouse IgE (BD Biosciences) at 2 μg/ml and antibody was detected with biotinylated anti-IgE (BD biosciences) at 0.1 μg/ml. Purified mouse IgE (BD Biosciences) was used as standard (starting concentration 500 ng/ml). For OVA-specific IgE detection, the plates were coated with anti-mouse IgE (BD Biosciences) as above and the antibody was detected by biotin-OVA (Nanocs) at 10 μg/ml. Mouse anti-OVA IgE (Bio-Rad was used as a standard (starting concentration 250 ng/ml). Antibody binding was detected with streptavidin-HRP (BD Biosciences) and developed with TMB substrate set (BD Biosciences).

Agaronyan K., Sharma L., Vaidyanathan B., Glenn K., Yu S., Annicelli C., Wiggen T.D., Penningroth M.R., Hunter R.C., Dela C.S, & Medzhitov R. (2022). Tissue remodeling by an opportunistic pathogen triggers allergic inflammation. Immunity, 55(5), 895-911.e10.

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