Measurement of the affinity of anti-D in the serum of immunized mothers and in immunoglobulin preparations with unlabeled antibodies.
BACKGROUND
Few data are available on the affinity of maternal anti-D responsible for hemolytic disease of the fetus and the newborn (HDN) and of anti-D used for the prophylaxis of that disease. A method was recently described to measure the affinity (K(a)) of untagged anti-D monoclonal antibodies (MoAbs). In this work, the same method was applied to determine the K(a) of polyclonal anti-D.


METHODS
O R(1)r red blood cells (RBCs) were sensitized with increasing concentrations of native anti-D in serum samples from immunized mothers and donors and in RhIG preparations. At equilibrium, the amount of anti-D bound to RBCs was measured by enzyme-linked immunosorbent assay. Scatchard and Langmuir equations were used to determine Ka.


RESULTS
The experimental data fitted well with the Scatchard equation (mean r2=0.95) but a better correlation was observed with the Langmuir equation (mean r2=0.99). The mean Ka of anti-D in 11 maternal serum samples, in 6 immunized donors, and in 5 lots of RhIG were 5.6x10(8) per M (from 2.8x10(8) to 12x10(8)/M), 3.9x10(8) per M (from 1.5x10(8) to 6.8x10(8)/M), and 3.4x10(8) per M (from 3.1x10(8) to 4.2x10(8)/M), respectively. The comparison of anti-D affinity in 5 cases of HDN with fetal anemia and in 6 cases of HDN with postnatal anemia showed no significant difference.


CONCLUSIONS
The method previously described for anti-D MoAbs was applied to polyclonal anti-D present in the serum of immunized subjects and in immunoglobulin preparations. The experimental data fitted well with the Langmuir equation, and the affinity of polyclonal of anti-D was measured with accuracy.
Debbia
Martine
M
Immunology Transfusion Unit, National Institute of Blood Transfusion, Paris, France.
Brossard
Yves
Y
Lambin
Patrick
P
D000906:Antibodies / Q000276:immunology*; D001782:Blood Donors; D001805:Blood Transfusion, Intrauterine; D004797:Enzyme-Linked Immunosorbent Assay; D004899:Erythroblastosis, Fetal / Q000097:blood / Q000175:diagnosis / Q000209:etiology / Q000276:immunology; D005260:Female; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood; D007136:Immunoglobulins / Q000097:blood*; D007231:Infant, Newborn; D007518:Isoantibodies / Q000097:blood*; D009035:Mothers; D011247:Pregnancy; D012204:Rh-Hr Blood-Group System / Q000276:immunology; D018029:Rho(D) Immune Globulin; D044967:Serum / Q000276:immunology*
D000906:Antibodies; D007074:Immunoglobulin G; D007136:Immunoglobulins; D007518:Isoantibodies; C061961:RHO(D) antibody; D012204:Rh-Hr Blood-Group System; D018029:Rho(D) Immune Globulin