In monoclonal antibody production, HAT medium is used to select for hybridomas by:

In monoclonal antibody production, HAT medium is used to select for hybridomas by:

• A. Allowing fusion of the two cell types
• B. Restricting the growth of myeloma cells
• C. Supporting growth of spleen cells only
• D. Blocking antibody production of IgM

Answer: (B)

HAT medium works by forcing cells to use the salvage pathway to make DNA. Aminopterin blocks the de novo synthesis route, so only cells that have functional HGPRT can survive. The myeloma cells used are HGPRT-deficient, so they cannot salvage nucleotides and die in HAT. Spleen cells, while not immortal, do have HGPRT and can survive briefly by salvage, but they’re not able to sustain growth long-term. When a spleen cell fuses with a myeloma cell, the hybridoma inherits HGPRT from the spleen cell and the immortal growth potential from the myeloma cell, allowing the fused cell to proliferate in HAT. This combination specifically restricts growth of unfused myeloma cells, enriching for the antibody-producing hybridomas.

HAT medium works by forcing cells to use the salvage pathway to make DNA. Aminopterin blocks the de novo synthesis route, so only cells that have functional HGPRT can survive. The myeloma cells used are HGPRT-deficient, so they cannot salvage nucleotides and die in HAT. Spleen cells, while not immortal, do have HGPRT and can survive briefly by salvage, but they’re not able to sustain growth long-term. When a spleen cell fuses with a myeloma cell, the hybridoma inherits HGPRT from the spleen cell and the immortal growth potential from the myeloma cell, allowing the fused cell to proliferate in HAT. This combination specifically restricts growth of unfused myeloma cells, enriching for the antibody-producing hybridomas.