Journal of Lipid Research, Vol 21, 902-912, Copyright © 1980 by Lipid Research, Inc.
Characterization and measurement of human apolipoprotein A-II by radioimmunoassay
RB Goldberg, JB Karlin, DJ Juhn, AM Scanu, C Edelstein and AH Rubenstein
The development of a radioimmunoassay for apolipoprotein A-II (apo A- II)
is described. Initial studies revealed a lack of immunological identity
between purified apo A-II used as the standard and serum or HDL. Extensive
testing of different buffers, standards, antisera, tracers, utilization of
a detergent, and heating of sera failed to resolve the problem. Gel
filtration of iodinated and non-iodinated apo A-II on Sephadex G-100
columns showed that apo A-II, in dilute solution, elutes in a higher
molecular zone than expected with a broad, assymetrical profile. The use of
a subfraction of the tracer in the assay resulted in parallelism in the
serum and standard dilution curves. The apo A-II assay was sensitive,
specific, and reproducible. Apo A-II added to sera was fully recovered and
delipidation did not affect the immunoreactivity of either serum or HDL.
Apo A-II contributed approximately 20% to the protein mass of HDL.
Comparison of these results with those obtained by radial immunodiffusion,
and with previously reported data, indicates that the reactivity of apo
A-II in its native and delipidated forms may be markedly influenced by
different immunologic methodologies and their specific reagents. Caution
should thus be shown at present in assigning absolute concentrations to apo
A-II in serum or HDL.
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