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Single B-cell Screening: The GEM Assay

We developed the GEM technology to interrogate the entire population of primary B cells from an immunized chicken. GEM stands for “Gel Encapsulated Microenvironment,” and it is our workhorse for identifying desirable antibodies in a large population of primary lymphocytes. The assay involves placing a single antibody-secreting lymphocyte in proximity with reporters (which can be cells or beads). The secreted antibody diffuses locally within the GEM and has the opportunity to bind to the reporters. Bound antibody can be detected either directly through the use of a secondary antibody, or by eliciting a response in the reporter that generates a visual signal. Each GEM may contain multiple types of reporters which can be differentiated from each other based on color.

Using The GEM Assay We Can:

1) screen up to 100 million cells in a single experiment

2) recover B cells directly from tissue without immortalization transformation or extended cell culture

3) facilitate multiparameter screening of antibody characteristics from a single B cell

4) directly identify biologically active antibodies

5) screen for antibodies binding with low nanomolar affinity or better

6) retain the heavy and light chain pairs created by in vivo affinity maturation

7) select 100 or more clones within a few hours after harvesting B cells from an immunized animal.

The image above shows a single GEM containing 2 types of reporter cells dyed with either a red or blue vital dye. Apoptotic cells are detected with a green Annexin V dye. Since green signal only co-localizes with blue signal in this experiment it can be determined that only the Type 2 reporter cells are sensitive to the treatment.

The GEM format has proven adaptable to a wide range of assays. The main assay requirement is that it produce a long-lived fluorescent signal. We have evaluated apoptosis, GPCR receptor signaling, specific binding to closely related proteins, binding to native conformation receptors on vesicles or on the surface of cells, staining of fixed tissue, and enzyme-blocking activity. For more details see our Publications page.