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#hybridoma cells

2 public questions tagged with this topic.

Hybridoma cells survive in HAT medium because they:

Survival in HAT environment demands functional purine and pyrimidine salvage enzymes inherited from normal B parent. B lymphocytes express HPRT1 locus on X chromosome encoding 24 kDa hypoxanthine-guanine phosphoribosyltransferase that catalyzes phosphoribosyl transfer from PRPP to hypoxanthine yielding IMP plus pyrophosphate and to guanine yielding GMP, bypassing de novo steps requiring folate. They also express cytosolic thymidine kinase 1 cell-cycle regulated peaking in S phase and mitochondrial kinase 2 constitutive, phosphorylating thymidine supplied in medium to TMP using ATP. Hybridoma c

Ref: Janeway Immunobiology salvage pathway HGPRT; Freshney Animal Cell Culture hybrids survive HAT via HGPRT TK from B cell.

Hybridoma cells survive in HAT medium because they:

Handling hybridoma cultures demands sensitive high-throughput detection secreted immunoglobulin microgram per mL among abundant serum proteins albumin bovine Ig. ELISA screening provides optimal format 96-well polystyrene plates coated purified antigen 1 ug/mL overnight 4 degrees passive hydrophobic adsorption van der Waals blocked 2 percent BSA 5 percent nonfat milk PBS preventing nonspecific binding plastic. Culture supernatant 50 uL added incubates 1h 37 degrees enabling specific binding Fab antigen interaction affinity association rate 10^5 per molar per second. After three washes PBS 0.05

Ref: Janeway Hybridoma Screening ELISA Technique Protocol; Alberts Protein A Purification Antibody Methods Chapter 8; Lodish Antibody Purification Immunotechniques Handbook Methods.