Complete saturation of protamine sulphate by dsDNA is necessary in order to obtain a highly sensitive and specific anti-dsDNA ELISA.
A protamine sulphate (PS) pretreated solid phase coated with different amounts of dsDNA has been used to develop a sensitive, specific and reproducible anti-dsDNA ELISA. Using low concentrations of a dsDNA coat 50% of SLE sera were found to be positive and false positive reactivity due to anti-PS reactivity was found in 3/40 patients with other auto-immune diseases (OAID). In contrast, when PS was saturated with higher concentrations of dsDNA 80% of SLE sera were detected, the reproducibility of the results was better and anti-PS reactivity of OAID patients with an anti-PS reactivity disappeared. The sera of three other OAID patients contained low avidity anti-dsDNA, measured after a salt elution step in the ELISA procedure, and 2/60 patients with non-auto-immune disease exhibited a false positive anti-dsDNA reactivity since they reacted with the solid phase even in the absence of PS and dsDNA. Thus an ELISA procedure using a PS pretreated solid phase permits the sensitive, specific and reproducible measurement of anti-dsDNA antibodies only if a high concentration of dsDNA is coated on the PS and appropriate controls are performed.
Rupin
A
A
Laboratory of Immunology, Faculté de Médecine, Tours, France.
de Jong
W
W
Degenne
D
D
Bardos
P
P
D000328:Adult; D000974:Antibodies, Antinuclear / Q000276:immunology*; D004247:DNA / Q000737:chemistry* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D006801:Humans; D066298:In Vitro Techniques; D008180:Lupus Erythematosus, Systemic / Q000276:immunology; D008297:Male; D008875:Middle Aged; D011479:Protamines / Q000737:chemistry*
D000974:Antibodies, Antinuclear; D011479:Protamines; D004247:DNA