The total IgM and IgG levels in B cell culture supernatants were measured with a Mouse IgM or a Mouse IgG ELISA Quantitation Set (Bethyl Laboratories, Montgomery, TX, USA), respectively, according to the manufacturer’s instructions. Two-fold dilutions of mouse reference IgM or IgG serum (Bethyl Laboratories) were used to generate standard curves. Sample dilutions giving OD readings in the linear portion of the appropriate standard curve were used to quantify the levels of Ig.
Mouse igg elisa quantitation set
Manufactured by Fortis Life Sciences
Sourced in United States
The Mouse IgG ELISA Quantitation Set is a laboratory equipment designed to quantitate the levels of mouse immunoglobulin G (IgG) in samples. It provides the necessary components to perform an enzyme-linked immunosorbent assay (ELISA) for the detection and measurement of mouse IgG.
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Lab products found in correlation
Mouse iga elisa quantitation set, by Fortis Life Sciences (3 mentions)
Mouse ifn γ elisa kit 2, by BD (1 mentions)
1 step ultra tmb elisa, by Thermo Fisher Scientific (1 mentions)
Maxisorp nunc immuno plate, by Thermo Fisher Scientific (1 mentions)
Maxisorp elisa plate, by Thermo Fisher Scientific (1 mentions)
Tmb substrate, by BD (1 mentions)
Anti mouse igg hrp, by Fortis Life Sciences (1 mentions)
Mouse cd19 microbeads, by Miltenyi Biotec (1 mentions)
El800, by Agilent Technologies (1 mentions)
Annexin 5 apoptosis detection kit, by BD (1 mentions)
Urea colorimetric assay kit, by Abcam (1 mentions)
Mouse igm elisa kit, by Fortis Life Sciences (1 mentions)
Microplate reader, by Agilent Technologies (1 mentions)
Mouse igm elisa quantitation set, by Fortis Life Sciences (1 mentions)
10 protocols using mouse igg elisa quantitation set
1
Measuring IgM and IgG Levels
2
Oral Immunization with Recombinant Salmonella Expressing TT
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Recombinant Salmonella (rSalmonella)–ToxC (ΔaroA, ΔaroD) and TT were kindly provided by the BIKEN Foundation (Osaka, Japan). Sox8−/− or littermate control mice were orally immunized with 5 × 107 CFU of rSalmonella-ToxC. TT-specific IgA and IgG in feces and serum were measured by ELISA. Flat-bottomed, 96-well MaxiSorp Nunc-Immuno plates were coated overnight with 500 ng/well of TT. Plates were blocked with 2% BSA in PBS, and optically diluted fecal extracts and sera were added into the plate wells. The Mouse IgA ELISA Quantitation Set (Bethyl Laboratories) and Mouse IgG ELISA Quantitation Set (Bethyl Laboratories) were used for antibody detection. To produce HRP signals, a 1-Step Ultra TMB-ELISA was used.
3
ELISA for T and B Cell Activation
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ELISA was carried out to evaluate T and B cell activation (Supplemental Fig. 3D ). After supernatants were harvested at day 5 in the T cell activation assay or at day 3 in the B cell activation assay, IFNγ and IgM were measured, respectively. Mouse IFNγ ELISA Kit II (BD Biosciences) and Mouse IgG ELISA Quantitation Set (Bethyl laboratories, Montgomery, TX) were used in this experiment according to the manufacturer’s instructions.
4
Salmonella Vaccine Immunogenicity in Mice
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rSalmonella-ToxC (ΔaroA, ΔaroD) and TT were kindly provided by the BIKEN Foundation (Osaka, Japan). Opg−/− mice and co-housed WT mice were orally or intraperitoneally infected with 5 × 107 or 5 × 105 colony-forming units (c.f.u.) of rSalmonella-ToxC1 (link),25 (link). TT-specific IgA and IgG in feces and serum were measured by ELISA. Flat-bottomed, 96-well Maxisorp Nunc-immuno plates (Thermo Fisher Scientific, Waltham, MA, USA) were coated overnight with 500 ng/well of TT. Plates were blocked with 2% BSA in PBS, and optically diluted fecal extracts and sera were added into the plate wells. The Mouse IgA ELISA Quantitation Set (Bethyl Laboratories, Inc., Montgomery, TX, USA.) and Mouse IgG ELISA Quantitation Set (Bethyl Laboratories, Inc.) were used for antibody detection. To produce horse radish peroxidase signals, a 1-Step Ultra TMB-ELISA (Thermo Fisher Scientific) was used.
5
Murine Stool IgA and IgG ELISA
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Murine stool (50 mg) was incubated in 500 μL of PBS at room temperature for 10 minutes, followed by vortexing for 5 minutes and bead-beating for 2 minutes. The sample supernatants were collected after centrifugation.
ELISA was performed using Mouse IgA ELISA Quantitation Set or Mouse IgG ELISA Quantitation Set (Bethyl Laboratories) according to the manufacturer’s protocol with minor modifications. MaxiSorp ELISA plates (Thermo Fisher Scientific) for total IgA measurement were pre-coated with 100 μL of diluted purified IgA antibody in 0.1M carbonate buffer (10 μl/mL). For OVA-specific IgA or IgG measurement, MaxiSorp ELISA plate were pre-coated with 5μg of OVA protein in 0.1M carbonate buffer. After overnight incubation at 4°C, plates were washed with PBS and incubated with PBS containing 1% BSA for 1 hour at room temperature. After washing, 100 μL of stool supernatant or serum were plated and incubated for 1 hour. Plates were washed again and 100 μL of anti-mouse IgA-HRP or anti-mouse IgG HRP (1:40 000 dilutions) was added before a 1-hour of incubation. TMB substrate (BD Pharmingen) was added to each well, followed by supplementation of 100 μL of 1M H2SO4 as a stop solution. Absorbance (450 nm) was measured using an ELISA reader.
ELISA was performed using Mouse IgA ELISA Quantitation Set or Mouse IgG ELISA Quantitation Set (Bethyl Laboratories) according to the manufacturer’s protocol with minor modifications. MaxiSorp ELISA plates (Thermo Fisher Scientific) for total IgA measurement were pre-coated with 100 μL of diluted purified IgA antibody in 0.1M carbonate buffer (10 μl/mL). For OVA-specific IgA or IgG measurement, MaxiSorp ELISA plate were pre-coated with 5μg of OVA protein in 0.1M carbonate buffer. After overnight incubation at 4°C, plates were washed with PBS and incubated with PBS containing 1% BSA for 1 hour at room temperature. After washing, 100 μL of stool supernatant or serum were plated and incubated for 1 hour. Plates were washed again and 100 μL of anti-mouse IgA-HRP or anti-mouse IgG HRP (1:40 000 dilutions) was added before a 1-hour of incubation. TMB substrate (BD Pharmingen) was added to each well, followed by supplementation of 100 μL of 1M H2SO4 as a stop solution. Absorbance (450 nm) was measured using an ELISA reader.
6
Isolation and Expansion of CD19+ TILs for IgG Detection
7
T Cell Modulation of B Cell Responses
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The wells of 96-well flat-bottomed plates were coated with
10 μg ml−1anti-CD3 mAb in 100 μl per well of PBS and incubated
overnight at 4 °C. The wells were washed, and MACS-purified
B cells with or without each FACS-purified T-cell subset (LAG3+ Treg,
CD25+ Treg
or CD4+CD25−CD44loCD62Lhi naive T cells)
or IL-27-treated CD4+ T cells described below were plated
immediately into the coated wells at a density of 1 ×
105 cells per well for each cell type in RPMI medium as
described above alone or with
10 μg ml−1anti-CD40 mAb
(3/23)+10 μg ml−1rIL-4 (Cell Signaling
Technology) supplemented with or without rTGF-β3
(1 ng ml−1). B cells
undergoing apoptosis on day 3 and total IgG production in the culture
supernatants on day 7 were determined using the Annexin V Apoptosis Detection Kit (BD
Pharmingen) and a mouse IgG ELISA
Quantitation Set (Bethyl Laboratories),
respectively, according to the manufacturer’s protocol.
10 μg ml−1anti-CD3 mAb in 100 μl per well of PBS and incubated
overnight at 4 °C. The wells were washed, and MACS-purified
B cells with or without each FACS-purified T-cell subset (LAG3+ Treg,
CD25+ Treg
or CD4+CD25−CD44loCD62Lhi naive T cells)
or IL-27-treated CD4+ T cells described below were plated
immediately into the coated wells at a density of 1 ×
105 cells per well for each cell type in RPMI medium as
described above alone or with
10 μg ml−1anti-CD40 mAb
(3/23)+10 μg ml−1rIL-4 (Cell Signaling
Technology) supplemented with or without rTGF-β3
(1 ng ml−1). B cells
undergoing apoptosis on day 3 and total IgG production in the culture
supernatants on day 7 were determined using the Annexin V Apoptosis Detection Kit (BD
Pharmingen) and a mouse IgG ELISA
Quantitation Set (Bethyl Laboratories),
respectively, according to the manufacturer’s protocol.
8
Serum Biomarkers in Murine Model
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Serum levels of antibodies to double stranded (ds) DNA (total IgA, IgG, and IgM) (Mouse anti-dsDNA antibodies total Ig ELISA Kit, Alpha Diagnostic, San Antonio, USA), IgM (Mouse IgM ELISA kit, Bethyl Laboratories, Montgomery, USA), IgG (Mouse IgG ELISA quantitation set, Bethyl Laboratories), interleukin-6 (IL-6) (Mouse IL-6 ELISA Kit, Life Technologies), urea (colorimetric Urea Assay Kit, Abcam, Cambridge, United Kingdom), C-terminal type I collagen (CTX-I, ELISA RatLaps kit, Immunodiagostic Systems, Boldon, UK), procollagen type I N propeptide (PINP, Immunodiagostic Systems), and parathyroid hormone (PTH, Elabscience, Houston, USA) were analyzed according to the manufacturers’ instructions.
9
IgM and IgG Detection in Murine Liver and Serum
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To detect the concentration of IgM and IgG in liver lysate of mice liver and serum, we used Mouse IgM ELISA Quantitation Set and Mouse IgG ELISA Quantitation Set (Bethyl Laboratories, USA). 80 μg liver lysate or 500 time diluted serum was performed ELISA analysis by following the standard protocol. OD450 was measured with a microplate reader (Bio-Tek, Winooski, VT).
10
IgG Quantification in Transgenic Mice
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The concentration of IgG in the plasma of 6–8 week old C57BL/6J, hFcRn Tg, and FcRn−/− mice was determined by ELISA. Blood samples were collected into heparinized tubes through submandibular cheek pouch bleeds. Blood was centrifuged at 6,000 rpm for 6 min and the plasma was stored at –80°C until use. Quantification of mouse IgG in plasma was determined using the Mouse IgG ELISA Quantitation Set (Bethyl Laboratories, Inc.; Montgomery, TX) following the manufactures recommended protocol. C57BL/6J plasma was diluted 1∶2500 in sample buffer (D-PBS, 0.05% Tween 20, 3% BSA) whereas hFcRn Tg and FcRn−/− plasma was diluted 1∶500 in sample buffer to achieve absorbance values within the range of the standard curve. A 1∶40000 dilution of the HRP detection antibody in sample buffer was used for this assay.
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