Identification and characterisation of cardiolipin autoantibodies

Herbert, Louisa (2003) Identification and characterisation of cardiolipin autoantibodies. Masters thesis, University of Central Lancashire.

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The aim of this study was to develop a method to isolate and identify anticardiolipin antibodies found in human serum samples obtained from patients with antiphospholipid syndrome (APS)
APS is a disorder with clinical manifestations that include venous and arterial thromboembolism, thrombocytopenia, recurrent spontaneous abortions, and possibly a variety of cardiac neurologic, dermatologic, and other problems, in association with the production of antibodies directed against phospholipids. Anticardiolipin antibodies are possibly the best studied and clinically most widely accepted antiphospholipid antibodies.
In this study affinity chromatography was optimised to facilitate the separation of antiphospholipid antibodies. Amberlite beads with a diameter of iooA were either used untreated as a control, or coated with cardiolipin in a column. Human serum was then passed through the column and eluted with three salt gradients (0.05-1 .OM NaCl)
It was also necessary to optimise an in house ELISA, for the measurement of ImmunoglobulinG and ImmunoglobulinM in the eluates. Finally Nuclear Magnetic Resonance (NMR) was used to investigate any possible chemical shift occurring when
cardiolipin was added to serum positive for high titres of anticardiolipin antibodies Results for IgG showed that although there was a slight increase in levels of IgG in positive serum samples after the salt gradients were added to the cardiolipin coated
column, a similar results was also seen in negative samples. However the control column did not show an elution of IgG for either positive or negative samples as expected. The results for 1gM were also inconclusive. The serum samples that were
positive for high titres of 1gM eluted antibodies after the salt gradient was added in the cardiolipin coated column, but also in the control column where it was expected that no further elution of 1gM was expected.
No results for NMR were obtained, as a standard could not be set. Overall, no definite conclusion can be made from the results as an elution of immunoglobulin was seen in both the control and cardiolipin coated column after the salt gradients had been added.

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