Immunogold labeling of the low-affinity (55 kd) IL2 receptor on the surface of IL2 receptor-bearing cultured cells and mitogen-activated peripheral blood lymphocytes

R. J. McGroarty, Gordon Mills, E. De Harven

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

Foreign antigens or mitogens initiate activation of T lymphocytes through antigen-specific receptors. After antigen-receptor interaction, T cells release a lymphotrophic hormone, interleukin 2 (IL2), and high-affinity IL2 receptors (IL2R) are progressively expressed on their surfaces. When evaluated under the scanning electron microscope after specific immunolabeling with colloid gold markers, the density of expression of the 55 kd low-affinity IL2 receptor (Tac) increases in a time-dependent manner, reaching a maximum 72 hours after the onset of phytohemagglutinin (PHA) activation. In contrast, the number of cells expressing the 55 kd molecule reaches a maximum by 24 hours and remains constant through 72 hours. Immunogold labeling allows simultaneous determination of IL2 receptor density at the level of each individual cell and the percentage of IL2 receptor-positive cells in any cell population under study. The rank order of receptor expression on the surface of established culture cell lines correlates with results reported from other laboratories with immunofluorescence and Scatchard analysis of binding of radiolabeled IL2. ALthough at high density of receptor expression immunogold labeling with 40 nm gold markers detects significantly fewer sites than radiolabeling, the method appears very sensitive in identifying low levels of expression. This results from 1) the extremely low non-specific background of the immunogold labeling technique and 2) the analysis of single cells rather than cell populations. Our results indicate that 2 cell lines, reported as negative for the expression of the 55 kd IL2 receptor (YT2C2 and MLA144), express low numbers of this molecule as demonstrated by immunogold labeling.

Original languageEnglish (US)
Pages (from-to)213-219
Number of pages7
JournalJournal of Leukocyte Biology
Volume48
Issue number3
StatePublished - Sep 12 1990
Externally publishedYes

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Interleukin-2 Receptors
Mitogens
Cultured Cells
Lymphocytes
Antigen Receptors
Interleukin-2
Single-Cell Analysis
T-Lymphocytes
Gold Colloid
Cell Line
Phytohemagglutinins
Gold
Population
Fluorescent Antibody Technique
Cell Count
Immunohistochemistry
Hormones
Electrons
Antigens

Keywords

  • colloidal gold
  • receptor expression
  • T lymphocytes

ASJC Scopus subject areas

  • Immunology
  • Cell Biology

Cite this

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title = "Immunogold labeling of the low-affinity (55 kd) IL2 receptor on the surface of IL2 receptor-bearing cultured cells and mitogen-activated peripheral blood lymphocytes",
abstract = "Foreign antigens or mitogens initiate activation of T lymphocytes through antigen-specific receptors. After antigen-receptor interaction, T cells release a lymphotrophic hormone, interleukin 2 (IL2), and high-affinity IL2 receptors (IL2R) are progressively expressed on their surfaces. When evaluated under the scanning electron microscope after specific immunolabeling with colloid gold markers, the density of expression of the 55 kd low-affinity IL2 receptor (Tac) increases in a time-dependent manner, reaching a maximum 72 hours after the onset of phytohemagglutinin (PHA) activation. In contrast, the number of cells expressing the 55 kd molecule reaches a maximum by 24 hours and remains constant through 72 hours. Immunogold labeling allows simultaneous determination of IL2 receptor density at the level of each individual cell and the percentage of IL2 receptor-positive cells in any cell population under study. The rank order of receptor expression on the surface of established culture cell lines correlates with results reported from other laboratories with immunofluorescence and Scatchard analysis of binding of radiolabeled IL2. ALthough at high density of receptor expression immunogold labeling with 40 nm gold markers detects significantly fewer sites than radiolabeling, the method appears very sensitive in identifying low levels of expression. This results from 1) the extremely low non-specific background of the immunogold labeling technique and 2) the analysis of single cells rather than cell populations. Our results indicate that 2 cell lines, reported as negative for the expression of the 55 kd IL2 receptor (YT2C2 and MLA144), express low numbers of this molecule as demonstrated by immunogold labeling.",
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AU - Mills, Gordon

AU - De Harven, E.

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N2 - Foreign antigens or mitogens initiate activation of T lymphocytes through antigen-specific receptors. After antigen-receptor interaction, T cells release a lymphotrophic hormone, interleukin 2 (IL2), and high-affinity IL2 receptors (IL2R) are progressively expressed on their surfaces. When evaluated under the scanning electron microscope after specific immunolabeling with colloid gold markers, the density of expression of the 55 kd low-affinity IL2 receptor (Tac) increases in a time-dependent manner, reaching a maximum 72 hours after the onset of phytohemagglutinin (PHA) activation. In contrast, the number of cells expressing the 55 kd molecule reaches a maximum by 24 hours and remains constant through 72 hours. Immunogold labeling allows simultaneous determination of IL2 receptor density at the level of each individual cell and the percentage of IL2 receptor-positive cells in any cell population under study. The rank order of receptor expression on the surface of established culture cell lines correlates with results reported from other laboratories with immunofluorescence and Scatchard analysis of binding of radiolabeled IL2. ALthough at high density of receptor expression immunogold labeling with 40 nm gold markers detects significantly fewer sites than radiolabeling, the method appears very sensitive in identifying low levels of expression. This results from 1) the extremely low non-specific background of the immunogold labeling technique and 2) the analysis of single cells rather than cell populations. Our results indicate that 2 cell lines, reported as negative for the expression of the 55 kd IL2 receptor (YT2C2 and MLA144), express low numbers of this molecule as demonstrated by immunogold labeling.

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