Hans Fey

Serological differentiation between Echinococcus granulosus and E. multilocularis infections in man.

An enzyme-linked immunosorbent assay (ELISA) was adapted for the serological differential diagnosis of cystic or alveolar echinococcosis in man caused byEchinococcus granulosus orE. multilocularis respectively. By affinity chromatography using rabbit anti hydatid fluid IgG coupled covalently to CNBr-Sepharose 4B a protein fraction (Em 1) containing shared antigens of both parasites could be isolated from an extract ofE. multilocularis metacestode tissue. From the same source another antigen fraction (Em 2) with a high degree of specificity forE. multilocularis was prepared by immunosorption. Antigen Em 1 was equally sensitive for the detection of antibodies againstE. granulosus andE. multilocularis, whereas antigen fraction Em 2 appeared to be more specific forE. multilocularis. A correct serological differential diagnosis was achieved in 95% of 57 confirmed cases of human cystic or alveolar echinococcosis by the simultaneous use of both antigen fractions in the ELISA and by comparison of their reactivities.

IMMUNOLOGY OF THE NEWBORN CALF: ITS RELATIONSHIP TO COLISEPTICEMIA*

As a result of these studies,l it has become generally accepted that specific agglutinating antibody against the K antigens of E . coli is the factor in colostrum that protects the calf against colisepticemia and other forms of colibacillosis. A colostrum without K-antibodies would be as worthless as no colostrum at alL4 Glantz and Jacks ZG also observed a positive correlation: Antibody in the serum was associated with survival of calves given E . coli serotype 115:K:H18, but not the less virulent 9:K35:T10. He only determined 0-antibodies. Gay and colleagues 22-24 do not share Brigs’s opinion. According to the former, it is most unlikely that K-agglutinins are the factor in colostrum protecting calves against colisepticemia, for it has been shown in field studies that although calves receive 0-agglutinin, in the colostrum they do not usually receive agglutinins against the K-antigens of the same E . coli strains. Calves fed colostrum and challenged with strain 137:K79:H41 were resistant to the colisepticemic form of colibacillosis regardless of the presence or absence of agglutinins to this strain in their sera. Vaccination of pregnant cows seems to be an effective means of controlling colibacillosis. Schoenaers and c011eagues,~~ in 172 herds, observed a decrease of calf mortality due to colibacillosis from 24.5% to 1.5%. In serum-protection experiments with strains 0 78 and 0 115 and sera with varying content of specific antibodies against‘ those two types, Dam lo saw a significantly higher rate of survival among calves treated with sera of high antibody content. examined a pool of some 30 colostrum sera serologically and by the mouse protection test and found a high antibody content against different pneumococcus types and E . rkusiopathiae, a moderate titer against strains of E . coIi and P . multocida, and a low titer against S. typkimuriuin and Sc. aronson. When absorbed by a given strain, the colostrum-serum pool still contained an amount of nonspecific protective substances against that given strain. The serum of the newborn calf either does not contain gammaglobulin or contains only traces of it, which do not appear in its serum until after the ingestion of colostrum, and the acquisition of antibodies is entirely dependent upon the acquisition of these globulins. The same conditions are present in the piglet, the lamb, kid, and foal. The site of absorption of these proteins is the Trainin

Immunological analysis of hemoglobin transition during metamorphosis of normal and isogenicXenopus

SummaryAntisera against larval and adultXenopus hemoglobins as well as adult human hemoglobin showed no cross-reaction when tested by immunodiffusion against each heterologous antigen. In this test hemoglobin of a single animal produced two precipitation lines for larvae, but only one for adult stages. Immunoelectrophoresis also revealed more complex precipitation patterns for larval than for adult hemoglobins. Hemoglobin of the isogenic hybrid cloneXenopus laevis/X. gilli also reacted with antisera against normalXenopus hemoglobin.Quantitation of hemoglobins, analyzed by radial immunodiffusion showed fewer than 1% of adult hemoglobin in red cells of larvae, but 30% at completion of metamorphosis. Two weeks later adult hemoglobin attained over 90%, and in red cells of adultXenopus an average of 1% larval hemoglobin were detected.The relatively short transition period suggests that the loss of larval hemoglobin may be due to the elimination of larval red cells, and that the increase in adult hemoglobin may be indicative of a new cell line.

Measurement of staphylococcal protein A and detection of protein A-carrying staphylococcus strains by a competitive ELISA method.

A competitive ELISA for the estimation of staphylococcal protein A is described. Tetanus toxoid is insolubilized on polystyrene and incubated with human antitoxin, which renders the Fc-piece of this antibody freely accessible to protein A. The binding of the latter is demonstrated by its competition with protein A-phosphatase conjugate. The method has been shown to be sensitive and reproducible. It has been used for the detection of protein A in culture supernatants and on living pathogenic staphylococci. The test could therefore be of diagnostic value. Protein A is also present in extracts of food contaminated with enterotoxic staphylococci. It can be eliminated by a simple absorption with insoluble porcine IgG.

A simple procedure for the production of Fab from bovine IgG as an absorbent in the preparation of class-specific anti-immunoglobulin

Abstract A simple and rapid procedure is proposed for the preparation of Fab from bovine IgG which is used as a single absorbent (removal of anti-light chain antibodies) in the production of class specific antisera anti γ (IgG), anti μ (IgM) and anti α (IgA). The papain digest is chromatographed on DEAE-cellulose equilibrated with 0·01 M PO 4 pH 8·0, then rechromatographed on DEAE-cellulose with 0·01 M Tris-HCl pH 8·6 + 0·05 M NaCl, then precipitated with ammonium sulfate at 60% saturation. The supernatant contains Fab which is free of the Fc piece and of intact IgG.

Eriochrome black, a means for reduction of nonspecificity in immunofluorescence.

Fluorescent preparations with a high degree of nonspecific background staining were treated with a 1 : 60 or 1 : 30 (w/v) aqueous solution of Eriochrome black for 10 sec and then remounted. Nonspecificity was thereby considerably reduced. The technique was applied to cryostat sections of guinea pig lymph nodes containing Serratia marcescens and E. coli, with tissues of mice suffering from a pneumococcus septicemia, with Brucella-positive bovine placentas and with brain impressions of rabid foxes, mice and a cat. In all the systems studied, the fluorescent conjugate was purified only by gel filtration through Sephadex G-50, a further fractionation by ion exchange chromatography was shown to be superfluous.

Electrophoretic comparison of cellular and soluble galactosyltransferase (lactose synthetase A protein) using specific antibodies.

Rabbit antisera against soluble human milk galactosyltransferase (GT) having anti-GT activity, as demonstrated by inhibition of enzyme activity were used for a comparative study of the molecular sizes of galactosyltransferase. For this purpose, affinity-purified antibodies were used for the identification of milk, serum and effusion galactosyltransferase from native or partially purified preparations resolved by sodium dodecyl sulfate polyacrylamide electrophoresis (SDS-PAGE) by the immune replica technique. Milk galactosyltransferase migrated as a 55-kilodalton (kD) protein, serum and effusion GT slightly faster. Cross-reactive enzyme forms of 110 kD and 20 kD were detected in milk only. In order to establish a relationship between intracellular and soluble galactosyltransferase, HeLa cells were metabolically labeled by [35S]-methionine, cells lysed, subjected to immunoprecipitation and the precipitate analyzed by SDS-PAGE/fluorography: a single band corresponding to the intracellular form of GT have similar mobility as the milk enzyme was detected. These results indicate a close structural similarity between soluble and cellular galactosyltransferase as judged by immunological cross-reactivity and electrophoretic mobility.

Use of a computer to evaluate sigmoidal curves in serology by a new procedure

Serological standard curves are mostly sigmoidal in shape. Their transformation into straight lines by linear regression can be the source of serious error. Log/log or logit/log handling of the values can straighten the curve but only if their distribution is normal. A new way of calculating concentrations of antibody or antigen which leaves the standard curve unmanipulated is described. Computer programs for TI 59 (Texas Instruments) and--in BASIC--for a personal computer have been written and greatly facilitate routine work.

Light chains, Fab and F(ab′)2 from bovine IgG: An easy method for their preparation as absorbents for class-specific anti-immunoglobulin

Abstract A very simple procedure is described for the preparation of light chains, F(ab′) 2 and Fab from bovine IgG. The reduced/alkylated substrate containing IgG, heavy and light chains is brought to neutrality, which precipitates the heavy chains. The remaining IgG is removed by ZnSO 4 treatment. The pepsin and papain digests are also submitted to ZnSO 4 precipitation which clears the supernatant from intact IgG and Fc fragments.

Improvement of the polyvalent Salmonella phage's O-1 diagnostic value by addition of a phage specific for the O groups E1–E4

The usefulness of the polyvalent Salmonella phage O-1 as a first step diagnostic tool is again emphasized. 96.1% of all Salmonella strains are lysed. Its disadvantage of not lysing some 10-30% of strains belonging to the O groups E1-E4 has been eliminated by creating a mixture of the phage O-1 with a phage G47, (obtained from Gershman) which is highly active on OE strains. Thus the mixture exhibits a high OE-specificity without impairment of the O-1 polyvalency.