For B1-8hi transfer experiments, C57BL/6 recipient mice were preimmunized by i.p. injection of 100 µg OVA (BioSearch Technologies) dissolved in PBS and precipitated in alum (Sigma-Aldrich) at a 1:1 ratio >2 wk before adoptive cell transfer. 12–24 h after B1-8hi cell transfer, recipient mice were immunized by i.p. injection of 100 µg NP19-OVA (BioSearch Technologies) precipitated in alum (Sigma-Aldrich) at a 1:1 ratio. For NP-Ficoll immunization, non-preimmunized C57BL/6 recipient mice were i.p. injected with 100 µg NP46-Ficoll (BioSearch Technologies) dissolved in PBS and precipitated in alum at a 1:1 ratio. For in vivo proliferation analysis, 1 mg EdU (Sigma-Aldrich) diluted in PBS was i.p. or i.v. injected at indicated time points after immunization. For PE immunization, mice were s.c. injected with 50 µl of an emulsion containing 50 µg of PE. Emulsions were prepared by mixing PE in PBS in a 1:1 ratio with complete Freund’s adjuvant (Sigma-Aldrich). For peptide immunizations, mice were i.p. injected with 100 µg Ag85b280-294 (FQDAYNAAGGHNAVF) or 2W1S (EAWGALANWAVDSA) peptide in 100 µl of a 1:1 emulsion with complete Freund’s adjuvant.
Np46 ficoll
Manufactured by Biosearch Technologies
NP46-Ficoll is a density gradient separation medium used for the isolation and purification of cells, organelles, and other biological particles. It is a sterile, endotoxin-tested solution of Ficoll polymer and a proprietary non-ionic polysaccharide that forms a continuous density gradient upon centrifugation. The core function of NP46-Ficoll is to facilitate the separation and enrichment of specific cell populations or subcellular components based on their density differences.
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Lab products found in correlation
2 protocols using np46 ficoll
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Preimmunization and Adoptive Transfer Protocol
2
Transgenic Mouse Strains for B Cell Study
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C57BL/6NCrSlc (B6) mice were purchased from Japan SLC. All the following mice were backcrossed to the B6 or B6-CD45.1 strain: B1-8flox/+ mice (Lam et al., 1997 (link)), B1-8hi mice (Shih et al., 2002 (link)), Igk–/– mice (Chen et al., 1993 (link)), Cd19cre/+ mice (Rickert et al., 1995 (link)), and Prkcd–/– mice (Miyamoto et al., 2002 (link)). Prkcd fl/fl mice on the B6 background were developed by Drs. Niino, Shioda, and Sakimura (Niino et al., 2021 (link)) and purchased from the RIKEN BioResource Center (RBRC06462). Mice were immunized i.p. with 100 µg of NP46-Ficoll (F-1420; Biosearch Technologies), 1 µg of PPS3 (169 X; American Type Culture Collection) or 100 µg of NP40-CGG in alum (Haniuda et al., 2016 (link)) unless otherwise noted. For flow cytometry of spleen cells, mice were immunized i.v. with 100 µg of NP46-Ficoll. Sex-matched 7–14-week-old mice were used for all experiments. All mice were maintained in the Tokyo University of Science (TUS) mouse facility under specific pathogen-free conditions. Mouse procedures were performed under protocols approved by the TUS Animal Care and Use Committee (Approval No. S19017 and S20011).
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