Radioautographic Studies of Poliovirus Binding by Human Immunoglobulins.

Abstract

A previous report from this laboratory demonstrated that radioactive poliovirus could diffuse through agar to combine with electrophoretically separated colostrum and serum antibodies ( 1 ) . The technique as used was relatively insensitive, requiring concentrated, purified immunoglobulins and large volumes of poliovirus. A more sensitive technique, combining immunoelectrophoresis and radioautography with radioactive antigen as one of the reactants, has been described for demonstrating antibody activity. Thus antibodies which bind insulin, thyroglobulins and ragweed have been detected in the IgG,t IgA and IgM classes of immunoglobulins by this method(2-6). The adaptation and use of this radioimmunoelectrophoresis technique to the study of antibody directed against poliovirus is described in this paper.Material and methods. Antigen. P2 labelled type I poliovirus was prepared by growing virus in HeLa cell cultures with citrate-Eagle's medium, 5 % dialysed calf serum and sodium phosphate P2 (0.066 mc/ml). The labelled virus was purified by ultracentrifugation and DEAE column chromatography ( 7 ) . The preparation used for diffusion contained approximately 10T.O TIDS0 and 100,-000 cpm per 0.1 ml.Serum. Serum specimens were obtained from adult human males and females who had not been immunized within the past 2 years. Serum polio neutralizing antibodies were demonstrated by a standard one hour tube-neutralization method, using 100 TIDBo of poliovirus, in HeLa cell cultures(8). The *This research was supponted by U.S.P.H.S. Research Grant AI-05009-03 from Nat. Inst. M t h .t The World Health Organization (Bull. 447, World Health Organ., v30, 1%4) has recommended the following nomenclature for the classes ob immumglobulins: G Immunoglobulin (IgG) for 7s (y2) -globulin ; A Immunoglobulin (IgA) for yla-globulin and M Immunoglobulin (IgM) for y,,,-globulin. Sinai Hospital, New York Citysera were fractionated by a combination of DEAE and Sephadex G-2Q0 column chromatography (9). The type of immunoglobulin in the eluates was determined by radial immunodiffusion( 10). The neutralization titer of each fraction was determined and correlated with the type of immunoglobulin present in the fraction.Radioimmunoelectrophoresis was performed as described by Yagi (2) and Morse(3) using agar consisting of 1% Difco Special Agar Noble in a 25% solution of 0.1 M Verona1 buffer (pH 8.6) on microscope slides. After electrophoresis of the serum, rabbit antiserum directed against human serum was added to the antiserum troughs. As soon as the antiserum had diffused into the agar, the radioactive poliovirus preparation was added to the trough. The plates were incubated 24 hours at room temperature and then washed for 48 hours with a 1% solution of NaCl. The slides were dried and then placed in contact with Kodak Royal Pan film for one week. Exposure was carried out in total darkness. After removal from contact with the film, the slides were stained with amido black. In some experiments the antiserum was allowed to react with the electrophoretic...

Cardiovascular control in diabetes mellitus.

“…of mean) in heart rate (0) and systolic blood pressure (Ci) measured 15 seconds, one minute, and five minutes after standing in (a) 13 patients (cases 1-13) with normal pattern of change in heart rate and systolic blood pressure on standing; (b) two patients (cases14 and 15) with initial fall in systolic blood pressure on standing and recovery without further increase in heart rate; (c) five patients (cases[16][17][18][19][20] with marked systolic hypotension and tachycardia on standing (P <0 001); and (d) one subject (case 21) with profound hypotension accompanied by only mild tachycardia.…”

Radioimmunoelectrophoretic identification of poliovirus inhibitors and their characteristic mode of action

“…Radioimmunoelectrophoretic identification of inhibitors in animal serum was performed using the procedures principally similar to those of AIlffBI~NDER et al (23). The active serum components were identified by comparing the radioautographie patterns with the corresponding immunoelectrophoregrams.…”