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#unfused cells

2 public questions tagged with this topic.

Immunological Applications

Myeloma cells used in hybridoma technology are selected to be:

Trastuzumab humanized IgG1 kappa mAb 148 kDa targets HER2 proto-oncogene extracellular domain IV juxtamembrane receptor overexpressed due ERBB2 amplification 17q12 20 percent breast gastric cancers poor prognosis. HER2 no known ligand but dimerizes HER1 HER2 HER3 after EGF binding autophosphorylating intracellular tyrosines Tyr1221 Tyr1222 recruiting Grb2 Shc activating PI3K Akt survival phosphorylating Bad Forkhead preventing apoptosis plus RAS RAF MEK ERK proliferation cyclin D1. Trastuzumab binds KD 5 nM sterically hindering dimerization preventing proteolytic cleavage shedding ECD generati

Ref: NEJM Trastuzumab HER2 Mechanism Slamon 2001 Clinical; FDA Herceptin Package Insert MOA Description; Janeway ADCC Trastuzumab Immune Mechanism NK Chapter 14.

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Immunological Applications

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.

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