Alkaline phosphatase conjugated goat anti mouse igm or igg antibodies

Serum immunoglobulin concentrations of various isotypes were analyzed by enzyme-linked immunosorbent assay (ELISA) [22 (link), 26 (link), 38 (link)]. Briefly, diluted sera were loaded onto precoated 96-well plates. Bound IgM or each IgG subtype was detected using alkaline phosphatase–conjugated goat anti-mouse IgM or IgG antibodies (SouthernBiotech) and an alkaline phosphatase substrate kit (Bio-Rad). Optical density at 450 nm was read on a microplate reader (BioTek Instruments).
The IgG anti–double-stranded DNA (anti-dsDNA) antibody-secreting cells (ASCs) were assessed by enzyme-linked immunospot (ELISpot) assay using MultiScreen filter plates (Millipore) as previously described [26 (link), 39 (link)]. Boiled salmon-sperm DNA (ThermoFisher Scientific)was used as the source of dsDNA.
Urinary protein concentrations were assayed using Uristix strips (Siemens Diagnostics).

Mouse sera antigen-specific antibodies were measured by ELISA. Plates were pre-coated with HA protein (1ug/ml), OVA (10 μg/ml), capture IgM or capture IgG antibodies. Mouse-specific antibody ELISA kits were used to measure IgM and IgG (Bethyl Laboratories, Montgomery, TX) as suggested by manufacturer. For ELISpot analysis, cells were incubated in HA-coated ELISpot plates (Millipore, Billerica, MA). Alkaline phosphatase-conjugated goat anti-mouse IgM or IgG antibodies (Southern Biotech, Birmingham, AL) were used as recommended by the manufacturer. ELISpot plates were developed using Vector AP substrate kit III (Vector Laboratories, Burlingame, CA) and quantified on a CTL plate reader and ImmunoSpot software (Cellular Technologies, Shaker Heights, OH).