Fusions & Subclonings

What cells are used in the fusion?

Splenocytes from mice with acceptable sera titers are fused with a murine myeloma cell line.

How are positive hybridomas selected?

Hybridoma supernatants are screened using a solid-phase ELISA; generally, with the same assay used to determine serum titers. Our criteria for a positive clone is signal twice the negative control.

Can I also screen at my lab?

Yes! We will send you the culture supernatants. The more data we have on antibody binding in different assays, the better the decision we can make selecting positive hybridomas.

Are positive hybridoma parent wells preserved, even if not selected for subcloning and production?

Cryopreservation of positive hybridomas is possible and should be discussed with your Project Manager. There is additional cost for freezing additional hybridomas.

Why do the positive hybridomas have to be subcloned?

Parent wells are mixtures of hybridoma clones. To isolate stable monoclonal lines we clone the parent well by limiting dilution. A second round of screening identifies positive, stable, cloned hybridomas. Two rounds of subcloning is the minimum to insure the cells are monoclonal.

What happens to positive subclones?

You and your Project Manager will decide which clones to expand, freeze, and put into the production phase. The selection is made on the basis of screening assay data. The more data from different assays that is available, the more ideal the selection of clones for expansion and/or freezing.

How long does it take to prepare clonal hybridoma cell lines?

Typically 4-6 weeks on cells from a single fusion. Some hybridomas are unstable and require more time.

How is pricing determined for these phases?

Pricing is set in our hybridoma development package but may vary if more clones are selected for cryopreservation or cloning, or if different assays are required. You receive a custom quote at the beginning of a project so your special needs can be addressed from the beginning, at a price you can afford.

 

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