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Improved plaque production for short-term bone marrow and fetal liver cultures
1Department of Biochemistry, Michigan State University, East Lansing 48824.
Abstract:
The addition of 0.5% globulin-free (GF-BSA) or 0.5% delipidated BSA (D-BSA) to short-term murine bone marrow (BM) (cultures) increased the number of plaque-forming cells (PFC) responding to trinitrophenylated lipopolysaccharide (TNP-LPS) 2-5-fold (1.1 X 10(4)-2.7 X 10(4) PFC per 10 X 10(6) nucleated BM cells). Although it was necessary to continue to supplement these cultures with 5% fetal calf serum (FCS), the inclusion of the aforementioned BSA preparations provided enhanced PFC production for all lots of FCS tested. Similarly, these preparations of BSA made it feasible to also culture BM in autologous mouse sera (MS) or in medium without 2-mercaptoethanol (2-ME) if in the latter case the D-BSA was pretreated with 2-ME. Thus, the inclusion of GF-BSA or D-BSA in short term cultures of BM not only substantially increased the number of Ig-secreting B cells produced in response to TNP-LPS but seemed to eliminate the need to screen for supportive batches of FCS or MS. These preparations of BSA also facilitated hapten specific PFC responses of fetal liver cultures.
Insights
Globulin-free (GF-BSA) and delipidated (D-BSA) bovine serum albumin preparations significantly enhance B-cell production in bone marrow cultures. These BSA additions improve plaque-forming cell responses to TNP-LPS, even with varied sera or reduced supplements.
Area of Science:
- Immunology
- Cell Biology
Background:
- Optimizing in vitro culture conditions is crucial for studying immune cell responses.
- Bovine serum albumin (BSA) is a common supplement in cell culture media.
- Specific BSA preparations may influence immune cell differentiation and function.
Purpose of the Study:
- To evaluate the effect of globulin-free (GF-BSA) and delipidated (D-BSA) BSA on murine bone marrow (BM) cultures.
- To determine if these BSA preparations enhance plaque-forming cell (PFC) production in response to trinitrophenylated lipopolysaccharide (TNP-LPS).
- To assess the utility of GF-BSA and D-BSA in supporting BM cultures with different sera or reduced supplements.
Main Methods:
- Murine bone marrow cells were cultured short-term with GF-BSA or D-BSA.
- Cultures were supplemented with fetal calf serum (FCS), mouse serum (MS), or serum-free medium.
- The number of PFC responding to TNP-LPS was quantified.
Main Results:
- Addition of 0.5% GF-BSA or D-BSA increased TNP-LPS-specific PFC numbers 2-5 fold.
- Enhanced PFC production was observed across various FCS lots.
- GF-BSA and D-BSA enabled successful BM culturing in autologous mouse sera or in medium lacking 2-mercaptoethanol (2-ME) under specific conditions.
- These BSA preparations also facilitated PFC responses in fetal liver cultures.
Conclusions:
- GF-BSA and D-BSA substantially increase Ig-secreting B cell numbers in short-term BM cultures stimulated with TNP-LPS.
- These BSA preparations reduce the need for screening supportive batches of FCS or MS.
- GF-BSA and D-BSA are valuable tools for enhancing immune cell culture and studying B-cell responses.