IgG and IgA antibodies were measured by enzyme‐linked immunosorbent assay (ELISA). On days 0, 7, 14, 21, 28, 35 and 42, mice were killed and samples harvested as previously described (Liu et al., 2014 ). Briefly, the plates were coated with purified lymphotoxin alpha (LTA) recombinant protein, lymphotoxin beta (LTB) recombinant protein, F41 fimbriae, 4 × STa recombinant protein and MBP‐STb recombinant protein at 50, 50, 200, 100 and 200 ng per well, respectively, then treated samples with serial dilution with PBS as the primary antibodies. HRP‐conjugated goat anti‐mouse IgA (Sigma‐Aldrich, St. Louis, MO, USA) or IgG (ZSGB‐BIO Co., Beijing, China) were used for bound antibodies detection as previously described (Jiang et al., 2014 ).
Hrp conjugated goat anti mouse iga
Manufactured by Merck Group
Sourced in France
HRP)-conjugated goat anti-mouse IgA is a laboratory reagent used in immunoassays. It consists of an antibody raised in goats against mouse immunoglobulin A (IgA), which is conjugated to the enzyme horseradish peroxidase (HRP). This reagent can be used to detect and quantify the presence of mouse IgA in various samples.
Automatically generated - may contain errors
Lab products found in correlation
Goat anti mouse iga, by Southern Biotech (1 mentions)
Sheep anti mouse iga, by Merck Group (1 mentions)
Goat anti mouse igg, by Southern Biotech (1 mentions)
3 3 5 5 tetramethylbenzidine (tmb), by BD (1 mentions)
Infinite m200 microplate reader, by Tecan (1 mentions)
Complete antiprotease cocktail, by Roche (1 mentions)
4 protocols using hrp conjugated goat anti mouse iga
1
Antibody Measurement Methodology
2
Quantification of Lipocalin-2 and IgA in Mouse Intestinal Samples
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Total lipocalin-2 concentrations of cecum content and fecal pellets were determined by sandwich ELISA using a commercial mouse lipocalin- 2/NGAL ELISA DuoSet (R&D, DY1857), according to the manufacturer’s instructions. Immunoglobulin A (IgA) concentrations were quantified from mouse intestinal lavages by sandwich ELISA. ELISA plates were coated with goat anti-mouse IgA (Southern Biotech, 1040-01) and IgA was detected with a horseradish peroxidase (HRP)-conjugated goat anti-mouse IgA (A4789, Sigma). A purified monoclonal IgA isotype antibody (Becton Dickinson, clone M18-254, 553476) served as standard. Absorbance was measured in a 96-microplate reader (VarioskanFlash, version 4.00.53) at 405 nm. Lipocalin-2 and IgA titers were analyzed in Prism 8 for Windows (GraphPad software Inc). −EC50 of each sample/standard was calculated by a four-parameter curve fitting.
3
Mouse Immunoglobulin ELISA Protocol
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Plates were coated with 5 µg/ml of sheep anti-mouse IgA (Sigma) or goat anti-mouse IgG (SouthernBiotech, Cliniscience, Nanterre, France), incubated with plasma, detected with 1.5 µg/ml HRP-conjugated goat anti-mouse IgA (Sigma) or goat anti-mouse IgG (SouthernBiotech). HRP was revealed using TMB and the reaction was stopped with H2SO4 before reading at 450 nm using Varioskan. microplate reader.
4
Quantification of Plasma and Fecal Immunoglobulins
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Intracardiac blood was recovered with heparined needle and plasma was kept at -80°C.
Fecal proteins were extracted mechanically in complete antiprotease cocktail (Roche Diagnostic, Meylan, France) and frozen at -80°C. France) before incubation with diluted samples or purified IgA or IgG (SoutherBiotech).
Plasma was diluted at 1:20,000 for IgG and fecal extracts at 1:6,000 for IgA. Horseradishperoxidase (HRP)-conjugated goat anti-mouse IgA (Sigma) or goat anti-mouse IgG (SoutherBiotech) were added, HRP was revealed using TMB (Becton Dickinson; BD, Le Pont de Claix, France) and the reaction was stopped with 25 µl of H 2 SO 4 2N before reading at 450 nm using an automatic Infinite M200 microplate reader (Tecan). Results were expressed as µg/ml for plasma and µg/mg of fecal proteins for IgA.
Fecal proteins were extracted mechanically in complete antiprotease cocktail (Roche Diagnostic, Meylan, France) and frozen at -80°C. France) before incubation with diluted samples or purified IgA or IgG (SoutherBiotech).
Plasma was diluted at 1:20,000 for IgG and fecal extracts at 1:6,000 for IgA. Horseradishperoxidase (HRP)-conjugated goat anti-mouse IgA (Sigma) or goat anti-mouse IgG (SoutherBiotech) were added, HRP was revealed using TMB (Becton Dickinson; BD, Le Pont de Claix, France) and the reaction was stopped with 25 µl of H 2 SO 4 2N before reading at 450 nm using an automatic Infinite M200 microplate reader (Tecan). Results were expressed as µg/ml for plasma and µg/mg of fecal proteins for IgA.
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