Details are given of a modified Euglena method for the assay of B12 in serum. Serum in very small quantities is usually inhibitory to the growth of the Euglena. In general, this inhibition is most marked when the organism is growing prolifically. For estimating serum B12 values, a standard curve is used from B12 standards containing 0·01 ml. of serum of low B12 content, so that growth in test serum solutions is compared with growth in serum-containing standards. The recovery of B12 added to serum, incomplete when estimated from aqueous standards, is now complete and the variation in results from batch to batch is markedly reduced.
Various factors—such as medium, light, and inoculum—can affect growth in aqueous and serum solutions differently and are an important cause of variations between batches and laboratories.
Optimal conditions were studied and they are essential for the most sensitive and accurate assay, but if the results are estimated from serum-containing standards accurate values can be obtained even under less than optimal conditions.
Subnormal serum B12 concentrations can be detected visually as early as 24 hours after the start of assay. For diagnostic purposes the assay can be read at three days, provided that appropriate dilutions are used, conditions are optimal, and growth is measured in a 1 cm. cell.
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