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PubMed1976

Modified Vi tests in the screening of typhoid carriers.

Two tests for the estimation of Vi antibody in sera were studied and their validity in the screening of typhoid carriers was assessed. One was a modified Vi haemagglutination test in which Vi-coated glutaraldehyde-fixed erythrocytes were used as the antigen and Vi antibody resistant to treatment with 2-mercaptoethanol was titrated. The other was a fluorescent Vi antibody test in which acetone-fixed bacterial Vi cells were used as the antigen and the total Vi antibody was titrated. In both tests, the antigens used were stable so that standardization of the tests would not be difficult. The modified Vi haemagglutination test was found equally sensitive but more specific in giving less false positives than the conventional Vi haemagglutination test. The fluorescent Vi antibody test was however found superior to both tests not only in giving less false positives but also in detecting more typhoid carriers.

Agglutination Tests↗
PubMed1976

Antibodies in human sera to oncorna virus-like proteins from normal or leukemia marrow cell cultures.

Some human marrows in culture release particles with oncornavirus-like properties. This study was designed to examine the immunological properties of similar particles in human marrow culture supernates. Leukemic and nonleukemic marrows were cultured for 5-7 days in the presence of [14C]uridine and [3H]leucine or [3H]glucosamine. Labeled supernatant components banding in sucrose gradient densities of 1.20-1.24 g/ml were used as antigen in a double antibody immunoprecipitation assay. The assay was validated by end point titrations and competition with unlabeled antigen; purified myeloma proteins were used as negative controls. Cross-reactivity with mammalian oncornaviruses, as judged by competitive inhibition of precipitation by these viruses, was slight and at the border of the sensitivity of the method. Precipitated antigens analyzed by SDS polyacrylamide gel electrophoresis contained three distinct polypeptides of about 70,000, 45,000 and 30,000 mol wt; these comigrated with the gp 70, pg 45, and p 30 of a murine leukemia virus. Similar polypeptides were obtained from both leukemic and nonleukemic marrow culture supernates. As determined by the radioimmunoprecipitation assay, 32 of 45 leukemic sera (71%), 36 of 45 normal sera (80%), 15 of 19 sera from family contacts of leukemic patients (79%), 14 of 21 cord blood specimens (67%), and 21 of 23 sera (91%) from patients with systemic lupus erythematosus had detectable antibody activity.

Antibodies, Viral↗
PubMed1978

Quantitation of antral gastrin cell populations in the dog.

A method for enumerating G cells in dog antral mucosa by a fluorescent antibody technique was shown to be valid with respect to specificity and reproducibility. The state of intracellular gastrin storage does not influence G cell identification by this technique. To obtain values representative of the entire antrum, all G cells in long strips of antral mucosa must be counted. Results are best expressed as G cells per centimeter length of mucosa. Even with large mucosal samples there are considerable variations in G cell density within a single antrum. The "average" dog antrum contains about 35 million G cells. We observed as much as 2-fold range in G cell density from one dog to another. Each animal should, therefore, serve as its own control when influences on G cell populations are to be studied experimentally.

Animals↗
PubMed1976

Immunization of mice and calves against Salmonella dublin with attenuated live and inactivated vaccines.

Previous findings, viz. that mice can be successfully immunized against infection with Salmonella dublin with either live or inactivated vaccine, were confirmed. Immunity lasted for at least 12 weeks in mice which had been immunized with inactivated alum-precipitated vaccine. The immunogenicity of inactivated vaccine gradually decreased on storage at 4 degrees C, but this was only detectable if a single injection was used for immunization: 2 injections virtually eliminated this phenomenon. The immunogenicity of live vaccine in mice was not enhanced by levamizole or the simultaneous injection of inactivated organisms. Both live and inactivated vaccines provided immunity in calves. A single injection of lyophilized vaccine, prepared from live rough Salmonella dublin strain (HB 1/17),protected 3 out of 6 calves, while 2 injections of a formalin-inactivated, alum-precipated vaccine, containing 1% packed cells of S. dublin strain 2652 V, protected 5 out of 6 calves against intraduodenal challenge with 2 x 10(9), S. dublin strain 2652 V. Two calves which had been immunized with an inactivated oil adjuvant vaccine were also solidly immune to this challenge. Serum antibody response in calves was poor when measured by the tube agglutination and the haemagglutination tests. Similarly, the sera had only marginal protective values when tested by means of a passive protection test in mice. Antibody titres alone are not a valid measure therefore, for the immune status of immunized animals.

Animals↗
PubMed1992

A supersensitive in-house enzyme-linked immunosorbent assay (ELISA) for measurement of thyroid-stimulating hormone (TSH) and its clinical applications.

A supersensitive ELISA was developed for measurement of thyroid-stimulating hormone (TSH) concentrations in serum using in-house rabbit polyclonal antisera and a commercial monoclonal antibody. The assay was optimised and validated by recovery, linearity and cross-reactivity experiments and further compared to other available assays and EQAS samples. Good precision was obtained with a working assay range of 0.2 to 100 mIU/L with < 10% coefficient of variation (CV) for both intra and interassay. The assay is highly sensitive and specific with a minimum detectable limit of 0.07 mIU/L and negligible cross-reactivities against LH, FSH, HCG and other pituitary peptides. Good correlations were obtained when compared to Abbott hTSH EIA (r = 0.993; p < 0.001; n = 85) and NETRIA IRMA (r + 0.995; p < 0.001; n = 76). The normal reference range established was 0.4 to 4.0 mIU/L (n = 76). TSH levels in serum of thyrotoxic patients (n = 83) were significantly lower (0.07 to 0.20 mIU/L, p < 0.0001) and completely distinct from normal values thereby obviating the requirement of a TRH-stimulation test. Stability studies showed that coated wells can be stored at 4 degrees C for at least 2 months. This highly sensitive in-house hTSH ELISA which is cheap, stable and readily available is useful for diagnosis and management of patients with various thyroid disorders.

Enzyme-Linked Immunosorbent Assay↗
PubMed1976

Does Freund's adjuvant denature protein antigens? EPR studies of emulsified hemoglobin.

Use of complete Freund's adjuvant for production of antibodies to study protein conformation is valid only if emulsification in adjuvant does not denature protein antigens. Using electron paramagnetic resonance to observe directly the protein in situ in the opaque emulsions, we demonstrate that hemoglobin is not denatured by emulsification or storage in adjuvant for 24 hr at 4 degrees C, conditions comparable to the usual handling of antigens before immunization.

Antigens↗
PubMedDate not supplied

[Clinical and biological alarm signs of drug allergy (author's transl)].

Allergic symptoms to medications, which cause treatment to be interrupted in order to prevent more serious accidents such as anaphylactic shock, are reviewed and their practical importance discussed. They are of small significance if functional as they can be produced by placebos, but are more disturbing if cutaneous and due to various non-specific and sometimes vague immuno-allergic mechanisms. Isolated fever of the allergic type has to be interpreted within the framework of the affection being treated. Raised blood eosinophil levels are a valid sign of an allergic antibody-type reaction. The value of detailed observation of symptoms and signs in such cases is demonstrated.

Antibody Formation↗
PubMed1976

Measurement of IgG-blocking antibodies: development and application of a radioimmunoassay.

A radioimmunoassay for measuring blocking antibodies has been developed. We used the ragweed antigen E system to show that the same blocking antibodies (IgG) measured by inhibition of antigen-induced leukocyte histamine release were precipitated in the binding assay (r8 = 0.96, p less than 0.001), thus validating a widely applicable technique for measuring blocking antibodies. Binding of phospholipase-A (Phos-A), the major allergen in honey bee venom, was also shown to correlate significantly with inhibition of histamine release. Hymenoptera (insect) hypersensitivity was used as a model to demonstrate application of the binding assay. Sera obtained from patients undergoing whole body extract therapy contained negligible amounts of specific blocking antibodies. Significantly higher blocking antibody titers to both whole honey bee venom and Phos-A were measured in sera drawn from patients immunized with whole venom. The use of the binding radioimmunoassay should facilitate management of allergic disease processes in which blocking antibodies are thought to be protective.

Antibodies↗
PubMed1979

Radioimmunoassay for immunologlobulin G in serum and urine.

We describe a radioimmunoassay for immunoglobulin G (IgG) in serum and urine. Aliquots of diluted samples and 125I-labeled IgG were incubated in antibody-coated tubes at 37 degrees C for 24 h, the supernates were decanted, and the radioactivity in tubes containing the bound fraction was counted. The dose-response curve in the range of 0.4--500 mg/L of urine or 640--40 000 mg/L of serum was linear on logit-log transformation and iterative weighted regression. Assay sensitivity was 10 ng of IgG. Validation studies included testing for precision, accuracy, antibody specificity, and parallelism of the dose-response curves for standard and unknown. In a study of 14 apparently normal individuals, serum IgG = 4.0-10.9 gL, urine IgG = 1.1-4.8 mg/24 h, and IgG clearance = 0.2 X 10(-4) to 4.8 X 10(-4) mL/min. In 20 patients with renal allografts, serum IgG = 15.8-66 g/L, urine IgG - 9.6-626 mg/24 h, and IgG clearance = 9 X 10(-4) to 1.99 X 10(-1) mL/min. IgG values correlated well with severity of renal allograft rejection.

Graft Rejection↗
PubMed1977

A direct radioimmunoassay for plasma renin in mice and its evaluation.

The pioneering work of Cohen, S., Taylor J.M., Murakami, K., Michelakis, A.M. and Inagami, T. ((1972) Biochemistry 11 4286-4293) with isolation of submaxillary renin and the first direct radioimmunoassay for renin subsequently described by Michelakis A.M. Yoshida H., Menzie, J., Murakami, K. and Inagami, T ((1974) Endocrinology 94, 1101-1105) was reinvestigated and confirmed. In addition a detailed evaluation of the assay is described by characterization of the immunoreactivity of the labelled renin and demonstration of immunological identity between plasma renin and purified renin standards. Assay and equilibrium conditions were characterized and optimized and the association constant for the antibodies determined to be of the order of 10(11)1/mol. The validity of the direct assay for renin was further investigated by a comparison with another radioimmunoassay for renin; the antibody trapping method, which measures the enzymatic activity of renin in plasma. The correlation coefficient is high (0.91) but does not exclude that the direct assay measures a hypothetical prorenin in plasma as well.

Animals↗
PubMed2026

Integrated Immunotherapy Target Atlas for Ewing Sarcoma.

BACKGROUND/AIM: Ewing sarcoma is a fusion-driven malignancy with low tumor mutational burden, making recurrent tumor-associated antigens with favorable tumor-to-normal contrast central to immunotherapy development. We converted the Deng et al.-defined 32-gene Ewing Sarcoma Specific Signature (ESS32) into a practical target atlas by integrating tumor RNA expression with normal-tissue context, protein evidence, subcellular localization, and therapeutic accessibility. MATERIALS AND METHODS: A 38-gene set was analyzed, including ESS32 and six comparator antigens (STEAP1, LINGO1, PRAME, CD99, CD276/B7-H3, and ENPP1). Eight Gene Expression Omnibus datasets (n=854 samples) were assigned predefined roles spanning tumor-versus-skeletal-muscle comparison, broad normal-organ context, EWSR1::FLI1 perturbation, tumor-only support cohorts, cell-line models, and cross-sarcoma comparison. Results were overlaid with Human Protein Atlas and published proteomic/surfaceome evidence. RESULTS: In GSE17674, the strongest tumor-enriched transcripts included NKX2-2, NPY1R, STEAP1, RBM11, RNF182, LIPI, CD99, STEAP2, LOXHD1, and DCDC2. Normal-tissue and compartment data substantially reordered RNA-only ranking. NKX2-2 showed the strongest Ewing-associated signal but encodes a nuclear transcription factor, favoring peptide-HLA/T-cell receptor (TCR) or vaccine development. RBM11 and LIPI emerged as high-interest intracellular/secretome-associated candidates, with an explicit epididymal/male reproductive caveat for LIPI. CD99 and NPY1R illustrated normal-cell reservoir and receptor-distribution constraints. CONCLUSION: ESS32 should be interpreted as an EWSR1::FLI1-associated RNA discovery set, not as a pre-validated target panel. Practical nomination requires integration of RNA enrichment, normal-tissue distribution, protein evidence, cellular compartment, and modality compatibility before nomination of TCR, vaccine, antibody-drug conjugate (ADC), chimeric antigen receptor (CAR), radioligand, or validation-first candidates.

Humans↗
PubMed1979

Localization of urinary tract infection.

Both the antibody-coating and LDH isoenzyme techniques theoretically fulfill the criteria for an ideal localization test. Both are noninvasive. They are relatively easy to perform and well within the capability of most clinical laboratories. Both can be done rapidly and the results can be in the clinician's hands in a matter of a few hours. The results of the antibody-coating technique do not appear to be valid for children. Since only one controlled study with comparison with other localization techniques has been done in adults, further evaluation needs to be done. The LDH isoenzyme pattern has not been studied in adults but appears to be accurate in children. A prospective comparison of both procedures in adults using either the Fairly bladder washout technique or the Stamey ureteral catheterization method to localize definitively the site of infection would be most helpful. Definitive noninvasive localization techniques could lead to therapy being initiated with the site of infection defined. This would facilitate earlier and more intensive antibiotic therapy in those patients at risk for renal parenchymal damage from upper tract infections.

Antibody-Coated Bacteria Test, Urinary↗
PubMed1979

Considerations for establishing the validity of immunocytological studies.

Immunocytology has wide spread applications for localizing tissue antigens, as evidenced by the recent exploitation of this technique in biological studies. Documenting the immunological specificity of the staining reaction is one of the most important technical considerations in validating the accrued data in immunocytological studies. The purpose of this report is to discuss and emphasize the need for conducting physiological studies in addition to the traditional immunological method and specificity controls. The ability of antibodies to bind molecules other than those molecules used as the immunizing material is a well documented fact. Hypothetically, preabsorption of the primary antibody with its specific antigen, could reduce subsequent binding of this antibody to a cross reactive tissue antigen, thus providing false confirmation of staining validity. The results of our experience with a cross reacting system in addition to other previously reported examples are discussed.

Animals↗
PubMed1992

Tenascin expression in basal cell carcinoma.

Tenascin (hexabrachion, cytotactin) is an extracellular matrix glycoprotein whose expression is strongly increased in hyperproliferative skin diseases, as shown by immunohistochemistry with polyclonal sera. In this study we describe a new monoclonal antibody (T2H5) against human tenascin. The specificity of T2H5 was validated by sequential immunoprecipitation of tenascin with polyclonal sera. T2H5 was used to analyse the presence of tenascin in basal cell carcinoma. Using Western blotting, at least two forms of tenascin were found, with approximate molecular weights of 210 and 300 kDa. In cultured human skin fibroblasts only the high molecular weight form was found.

Antibodies, Monoclonal↗
PubMed1975

[Gastrin].

Gastrin is released by food rich in proteins and by vagal mechanisms. HCI and possibly secretin and glucagon inhibit gastrin release. In the wide range of actions of gastrin, stimulation of gastric acid secretion is the most important. With the advent of radioimmunochemical methods for the determination of gastrinaemia, it has been shown that gastrin exists in a number of forms of different molecular weight. To estimate the validity of gastrin radioimmunoassay it is necessary to demonstrate that decrease in antibody-bound labelled antigen is unrelated to non-specific interference by unknown substances present in serum samples, and that the antiserum reacts with endogenous hormone in an identical manner. Heterogeneity of gastrin in serum may affect the validity of the radioimmunoassay. Hypergastrinaemia associated with hyper-normochlorhydria occures in gastrinoma, hyperplasia of antral gastrin cells, diseases with delayed gastric emptying, retained antrum, short bowel syndrome,renal failure. Hypergastrinaemia associated with hypo-achlorhydria occurs in atrophic gastritis without extensive antral lesion and after vagotomy. Gastrin radioimmunoassay can be used for the mass screening of subjects with atrophic gastritis, a high risk group for gastric cancer.

Antibody Formation↗
PubMed1992

[The factors that affect the results of the serological diagnosis of HIV infection and the screening for anti-HIV].

The laboratory diagnosis of HIV infection and screening for anti-HIV have determined the factors that influence the results of studies performed by ELISA and Western immunoblot. The factors include sensitivity and specificity of the test systems used, storage conditions of and transportation conditions for serum, errors of procedures for reactions, etc. Correlation is between the inactivation of the sera studied and the initial antibody titers during their storage at elevated temperatures. To obtain more valid results of HIV infection serodiagnosis it is necessary to use some test systems differing in the principles of setting up a reaction.

AIDS Serodiagnosis↗