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Flow Injection Analysis for Simultaneous Quantification of Prolactin Concentration and Glycosylation Macroheterogeneity in Cell Culture Samples

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Abstract

A flow injection analysis (FIA) system is presented for a twostep immunoassay-based determination of the total humanprolactin (hPRL) concentration along with its degree ofglycosylation. Separate measurement of total hPRL and nonglysosylated human prolactin (nG-hPRL) were made using twoflow-through cartridges each containing immobilized antibodiesof different specificity. The antibodies are immobilized on thesurface of a carrier. Glycosylated hPRL (G-hPRL) and, thus, thedegree of glycosylation were calculated by the differencebetween the two specific determinations. Enhanced specificityfor the determination of nG-hPRL was obtained using unfavorablebinding conditions through incorporation of alkaline pH andchaotropic agents into the carrier/dispersion buffer. The assayfor total hPRL and nG-hPRL were each found to be linear withinthe relevant concentration range. The results of the two-stepFIA method were found to agree with those obtained by thestandard methods of ELISA and western blotting while offeringthe advantage of minimal analysis time (10 min) and eliminationof manual manipulations.

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Authors and Affiliations

  1. Department of Chemical Engineering and Biotechnology Process Engineering Center, Massachusetts Institute of Technology, Room 56-469, Cambridge, MA, USA

    Martin Reinecke

  2. Department of Chemical Engineering and Biotechnology Process Engineering Center, Massachusetts Institute of Technology, Room 56-469, Cambridge, MA, USA

    Gregory Stephanopoulos

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  1. Martin Reinecke
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  2. Gregory Stephanopoulos
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Correspondence to Gregory Stephanopoulos.

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Reinecke, M., Stephanopoulos, G. Flow Injection Analysis for Simultaneous Quantification of Prolactin Concentration and Glycosylation Macroheterogeneity in Cell Culture Samples. Cytotechnology 34, 237–242 (2000). https://doi.org/10.1023/A:1008170023099

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