The fused cells were cultured in 1 × hypoxanthine-aminopterin-thymidine culture medium (H0262, Sigma, St Louis, MO, USA). Cells displaying a positive ELISA test signal (see below) were transferred to a 24-well plate. After separating the cells into 96-well plates, they were cultured for 7–10 days in hypoxanthine and thymidine culture medium (11067-030, Gibco, Carlsbad, CA, USA) under 5% CO2 at 37 °C in an incubator. Hybridoma cells were screened using ELISA (see below) and the cloning process was repeated until the final antibody secreting clone was selected.
Hypoxanthine aminopterin thymidine culture medium
Hypoxanthine-aminopterin-thymidine (HAT) culture medium is a specialized growth medium used in cell culture applications. It is formulated to support the selective growth of cells that have undergone genetic modification or hybridization. The medium contains hypoxanthine, aminopterin, and thymidine, which together inhibit the de novo synthesis of nucleotides, thereby selecting for cells that rely on the salvage pathway for DNA synthesis.
2 protocols using hypoxanthine aminopterin thymidine culture medium
Generating Anti-Citrullinated Peptide Antibodies in Mice
The fused cells were cultured in 1 × hypoxanthine-aminopterin-thymidine culture medium (H0262, Sigma, St Louis, MO, USA). Cells displaying a positive ELISA test signal (see below) were transferred to a 24-well plate. After separating the cells into 96-well plates, they were cultured for 7–10 days in hypoxanthine and thymidine culture medium (11067-030, Gibco, Carlsbad, CA, USA) under 5% CO2 at 37 °C in an incubator. Hybridoma cells were screened using ELISA (see below) and the cloning process was repeated until the final antibody secreting clone was selected.
Monoclonal Antibody Generation Against Citrullinated Peptide
The fused cells were then cultured in hypoxanthine–aminopterin–thymidine culture medium (Sigma, St. Louis, MO, USA). Cells showing a positive signal in the ELISA were transferred to a 24-well plate. After individual cells were placed into separate wells in 96-well plates, the cells were cultured for 7–10 days in hypoxanthine and thymidine culture medium (Gibco/Thermo Fisher Scientific, Waltham, MA, USA) in a 5% CO2 incubator at 37°C. Hybridoma cells were screened by ELISA, and the cloning process was repeated until the final antibody-secreting clone was selected.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!