ENUMERATION OF HUMAN PERIPHERAL-BLOOD LYMPHOCYTES SECRETING IMMUNOGLOBULINS OF MAJOR CLASSES AND SUBCLASSES IN HEALTHY-CHILDREN AND ADULTS

The reverse enzyme-linked immunospot assay was modified to enumerate peripheral blood mononuclear cells (PBMC) secreting IgG1-4, IgA1-2, and IgM. Anti-human IgG F(ab')2 and mouse monoclonal antibodies specific to each of the isotypes were used as solid-phase capture antibodies and developing antibodies, respectively. Although attempts were also made to detect IgD- and IgE-secreting cells (SC), their numbers in the peripheral blood were too few to be reliably estimated. The assay was applied to study healthy subjects including 21 neonates within 3 days of birth, 44 1- to 48-month-old children, and 32 adults. Sixty percent of these neonates had IgM SC in small numbers (< 20 per 10(6) PBMC), but only three neonates had IgSC of other isotypes. In contrast, by 1-2 months of age children had IgSC of all isotypes, including IgA2 and IgG4, often in higher numbers than adults. The relative frequencies of IgSC were IgA1 > IgG1 > IgM > IgG2 > IgG3 > IgG4 > IgA2 in the children and IgA1 > IgG1 > IgA2 > IgM > IgG4 > IgG2 > IgG3 in the adults. The order of the serum concentrations in the adults was IgG1 > IgG2 > IgA > IgM > IgG4 > IgG3. No correlation was found between the serum level and the IgSC number of individual isotypes (except for serum IgA and IgA1-SC). This new methodology should facilitate investigating the current status of immunoglobulin synthesis and the Ig repertoire in adults and children, in health and in disease.