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Simultaneous Expression of Hapten Binding and Antibody Secretion by NIP-Primed Lymph Node Cells Early in the Immune Response¹

From the Department of Pathobiology, University of Pennsylvania School of Veterinary Medicine, Philadelphia, Pennsylvania 19104

Abstract

A combination rosette-plaque assay was used to demonstrate that most (92%) of the plaque-forming cells (PFC) bound hapten to their surface 5 to 8 days after one injection of 3-nitro-4-hydroxy-5-iodophenyl acetic acid (NIP) coupled to diphtheria toxoid, (NIP-TOX). There was a significant decrease in hapten binding by these cells later in the primary response. Rosette formation was observed when NIP-TOX-primed lymph node cells bound NIP-coupled sheep red blood cells stabilized with pyruvic aldehyde (NIP/PASRBC). This suspension of cells was then plaqued on fresh NIP-coupled sheep red blood cells (NIP/SRBC). A rosetted lymph node cell in the center of a plaque clearly demonstrated simultaneous hapten binding and anti-hapten antibody secretion. The specificity of hapten binding and antibody secretion was established by rosette and plaque inhibition with NIP coupled to bovine serum albumin (NIP-BSA). The specificity of each was further characterized by comparative inhibition with analogues of NIP (also coupled to BSA): 3,5-dinitro-4-hydroxyphenyl acetic acid (NNP-BSA), 3,5-diiodo-4-hydroxyphenyl acetic acid (DIP-BSA) and 3-nitro-4-hydroxyphenyl acetic acid (NP-BSA). The homologous hapten NIP was always the best inhibitor of plaques (97.8% inhibition) and rosettes (96.2% inhibition). NNP was always a better plaque and rosette inhibitor than either DIP or NP. This hierarchy of inhibition, and the correlation between rosette and plaque inhibition demonstrated the similarity of receptor and antibody specificity.

Footnotes

¹ This research was supported by National Science Foundation Grant GB 28694, National Institutes of Health Training Grant TOICAO5097, and National Institutes of Health Career Development Award 1OKA16164 to B.W.