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Anal Chem. 2001 Sep 1;73(17):4333-7.

Assay of spherical cell surface molecules by thermal lens microscopy and its application to blood cell substances.

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Department of Forensic Medicine, Juntendo University School of Medicine, Hongo, Tokyo, Japan.


To detect and quantitate uneven cell surface molecules, such as blood group antigens on a blood cell and immunoglobulin molecules on a mast cell, an improved method of thermal lens microscopy was employed. The antigen molecules were immunologically stained with their antibodies, which were labeled with colloidal gold. Since the surface of the biological cells was not flat but spherical, the focal point of the probe laser beam inevitably deviated from the sample surface on the moving stage. Therefore, the deviation of the focal point of the probe beam was corrected by adjusting the phase of the signal. Using this technique, a three-dimensional antigen distribution on each cell surface was imaged. Despite the convex surface of cells, labeled colloidal gold was correctly quantified. In the measurement of erythrocyte antigens, a small quantity of Lewis antigens was successfully detected on the umbilical cord erythrocytes. Immunoglobulin E on a mast cell, derived from the allergic human mucosa fungus, was also observed by this method, and the distribution of IgE molecules on the cell surface was quantitatively imaged. A thermal lens microscope, which measures spherical samples correcting the deviation, made it possible for us to observe and assay the substances on biological specimens that have complicated forms, such as living cells in vivo or in situ.

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