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PubMed1975

Immunochemical comparison of estradiol-binding molecules in perinatal rat brain cytosol and serum.

Estradiol-binding macromolecules in fetoneonatal rat brain cytosol and serum were compared by immunochemical techniques. When treated by a double diffusion procedure, both cytosol and serum formed precipitin lines with rabbit antiserum specific for perinatal rat serum proteins. These lines fused completely, indicating, within the limits of detection of this particular antiserum, the presence of identical antigenic determinants in the brain and serum. Prior removal of immunoprecipitable material from cytosol or serum, after incubation with the specific antiserum, prevented formation of such precipitin lines. The procedure similarly presented specific estradiol-binding to macromolecules. It was therefore concluded that the specifically perinatal, antigenically similar, components in rat brain cytosol and serum (possibly representing alphafetoprotein) are responsible for the estradiol-binding activity in these two tissue compartments. Measurements of heme concentrations indicated that the alphafetoprotein-like material in the cytosol does not reflect blood contamination, but rather a separate population of similar or identical molecules.

Animals↗
PubMed1981

Glomerular disease in two patients with urticaria-cutaneous vasculitis and hypocomplementemia.

We assessed the clinical, serologic, and renal abnormalities of two patients with the syndrome of hypocomplementemia, vasculitis, and urticaria. Both patients presented with recurrent urticaria and were found subsequently to have glomerulonephritis. Both manifested depletion of the early complement (C) components, and their Cl inhibitor levels were normal. Circulating immune complexes were detected by two of three assays in both patients. One patient had a high titer of Sm antibodies. The detectable immune complexes were of high molecular weight, and were more suggestive of classical lupus erythematosus than the low molecular weight (7S) Clq precipitins which characterize some cases of hypocomplementemia-vasculitis-urticaria syndrome. Biopsy of the skin lesions in both patients revealed leukocytoclastic vasculitis with immunoglobulin and C deposits at the dermal-epidermal junction. Renal biopsy specimens showed segmental or diffuse proliferative mesangial glomerulonephritis. The results suggest that patients with this syndrome do not conform to a discrete entity but rather exhibit a spectrum of diseases ranging from systemic lupus erythematosus to the syndrome of urticaria, hypocomplementemia, and low molecular weight Clq precipitins.

Adult↗
PubMed1977

Aspects of the humoral response to laboratory white mice infected with the metacestodes of Taenia crassiceps.

Two strains of laboratory white mice, CFLP and LACA, were infected with the metacestodes of Taenia crassiceps. The serum from each infected mouse was tested for antibodies on double-diffusion gel plates and the number of antigen-antibody precipitin lines formed related (a) to the duration of the infection and (b) to the volume of metacestodes. There was a significant increase in the number of antigen-antibody precipitins with regard to the time infected but not to the volume of metacestodes. The pattern of development of the antigen-antibody precipitation system was also examined.

Animals↗
PubMed2000

Diagnosis of allergic bronchopulmonary aspergillosis (ABPA) in cystic fibrosis.

BACKGROUND: The diagnosis of allergic bronchopulmonary aspergillosis (ABPA) in cystic fibrosis (CF) patients may be difficult to establish because ABPA shares many characteristics with coexisting atopy or other lung infections in these patients. This study aimed to evaluate the sensitivity and specificity of various paraclinical parameters in the diagnosis of ABPA in patients with CF. METHODS: Accumulated data from a 5-year period in 238 CF patients were used to divide patients into two groups designated the ABPA group (n=26) and the non-ABPA group (n = 35). Patients in both groups were colonized with Aspergillus fumigatus (Af.), but only the ABPA group consistently demonstrated specific IgE antibodies and specific precipitins. Patients without A. fumigatus colonization were not assigned to either of these groups (n = 177). By this selection as the true diagnosis, 10 patients were selected from the ABPA group and 10 patients from the non-ABPA group. RESULTS: The groups were comparable as to age, sex, lung function (P=0.6), and presence of chronic Pseudomonas aeruginosa infection (P>0.1). No significant difference between the groups in unspecific atopic parameters such as eosinophil count (P=0.9) or eosinophil cationic protein (ECP) in sputum, plasma, or serum (P=0.9, P=0.59, and P = 0.9, respectively) was demonstrated. Total IgE was significantly higher in the ABPA group (P<0.01). The groups were comparable in skin prick test (SPT) positivity to a standard panel of aeroallergens (pollen, dander, molds, and mites) (P>0.2). Statistically significantly higher levels in the ABPA group were demonstrated in specific IgE to Af. (P < 0.05), SPT positivity to Af. (P < 0.02), and Af. precipitins (P < 0.05). Histamine release (HR) to Af. tended to be higher (P=0.075) in the ABPA group. Specific IgE to Af. was determined by Magic Lite (ML), CAP, and Maxisorp (in-house RAST). The CAP level was one to two classes higher than the ML level; however, the results were comparable (r=0.66, P<0.005). IgE to Af. measured by CAP was the test which offered the highest positive predictive value (PPV) and negative predictive value (NPV). Optimal diagnostic cutoff levels for the diagnosis of ABPA were determined: class 2 for HR to Af., 200 kIU/l for total IgE, and 3.5 (titer) for precipitating antibodies to Af., and class 2 for IgE to Af. (by CAP System). CONCLUSIONS: Unspecific atopy markers were of limited value for the diagnosis of ABPA. Patients with ABPA do not seem to be more atopic to other aeroallergens than non-ABPA patients. The most valid parameters for the diagnosis of ABPA in CF are SPT to Af., IgE to Af. in combination with precipitating antibodies to Af., and/or total IgE.

Adolescent↗
PubMed1975

Extraction and preliminary characterization of a human bronchogenic carcinoma antigen.

Saline extracts of human bronchogenic tumours, soluble in 50% saturated ammonium sulphate and also fractions from Sephadex G-200 chromatography were used to raise antisera in rabbits. After absorbing the antisera with normal tissue extracts, direct Ouchterlony tests were performed against tumour (adenocarcinomata and squamous cell carcinomata) and normal extracts. A precipitin reaction was given with all 11 tumour extracts tested at a concentration of 5 mg/ml whereas all the 9 normal lung control extracts did not react at concentrations up to 100 mg/ml. The possibility that this reaction could be related to histocompatibility differences between individuals is ruled out by the fact that in two cases tumour and normal tissue were obtained from the same patient. These studies and also precipitin-inhibition experiments have confirmed the existence of antigen associated with bronchial carcinomata and have shown that, although the antigen or a cross-reacting antigen is present in normal lung tissue, the amounts are small in comparison with the amounts extracted from tumour. Antigenic activity was contained in a single absorbance peak when fractionated by Sephadex G-200 chromatography and its elution volume indicated a molecular weight of approximately 4-0 times 10(4)D. Further purification was achieved using isotachophoresis. Preliminary characterization of the antigen has shown it to be stable at pH 4-5, resistant to heating at 50 degrees C for 30 min, to migrate on immunoelectrophoresis with a cationic mobility at PH 8-5 and to be immunologically distinct from carcinoembryonic antigen.

Adenocarcinoma↗
PubMed1981

Studies on the thermal degradation of the H and M antigens of lyophilized histoplasmin.

Lyophilized histoplasmin for the agar gel microimmunodiffusion test has been prepared as a candidate World Health Organization Biological Reference Reagent. It was subjected to elevated temperature for given periods of time and analyzed by the capillary precipitin test and the single radial immunodiffusion test to determine the stability of the H and M antigens. H antigen showed no fall in relative potency when incubated at 48 degrees C for 20 days. M antigen showed a fall in relative potency after storage at 37 degrees C and 48 degrees C, but the extent of the fall was greater in the radial immunodiffusion test than in the capillary precipitin test. Half-lives of the antigens could not be calculated from the Arrhenius equation because the response curves at each temperature followed different kinetics. However, as based on zero time data, M antigen of the lyophilized histoplasma showed a 20% drop in relative potency when stored at -20 degrees C for 2 years. Other analyses suggested that M antigen of liquid histoplasmin stored at 5 degrees C and of lyophilized histoplasma stored at -20 degrees C was degraded at equal rates.

Animals↗
PubMed1966

Studies on the effect of the carrier molecule on antihapten antibody synthesis. I. Effect of carrier on the nature of the antibody synthesized.

1. Fluorescence quenching has been calibrated, by comparison with equilibrium dialysis, for measurement of antibody-hapten association constants with guinea pig antibody to the 2,4-dinitrophenyl (DNP) determinant. The maximum quenching with all sites occupied which gave best agreement with equilibrium dialysis was found to be 100%. 2. The carrier to which the DNP-hapten is coupled influences significantly the amount, class (gamma(1) or gamma(2)), and affinity of the antibody of DNP specificity made by individual guinea pigs. 3. The affinities of gamma(1)- and gamma(2)-antibodies from individual guinea pigs are generally very similar. 4. Precipitin curves with gamma(1)-antibodies show more marked inhibition of precipitation in antigen excess than do precipitin curves with the less charged gamma(2)-antibodies, indicating the importance of nonspecific forces in the precipitation reactions.

Aminocaproates↗
PubMed1966

Studies on the immunochemistry of streptococcal mucopeptide.

Streptococcal mucopeptide, solubilized by either ultrasonic treatment or lysozyme, gave a precipitin reaction with rabbit antimucopeptide serum. A haptenic inhibitor of this reaction, which was composed of alanine, glutamic acid, and lysine in a mole ratio of 4:1:1, was isolated from a Streptomyces albus enzymes digest of Group D cell walls by ion exchange chromatography. When selected antisera were employed, greater than 90% inhibition of the mucopeptide quantitative precipitin reaction was achieved with 2 mg/ml of this inhibitor, whereas a hexosamine fraction with minimal concentrations of amino acid residues was inactive in this respect. These results suggest that the peptide moiety is an antigenic determinant of mucopeptide. Preliminary results indicate that the hexosamine polymer of the mucopeptide is a secondary antigenic determinant.

Antigen-Antibody Reactions↗
PubMed1981

Immunochemical specificity of the combining site of murine myeloma protein CAL20 TEPC1035 reactive with dextrans.

The immunochemical specificity of the combining sites of murine myeloma protein CAL20 TEPC1035 was studied by quantitative precipitin and precipitin inhibition assays. Myeloma protein CAL20 TEPC1035 precipitated with only three dextrans, B1355S4, B1498S, and B1501S, with high proportions of alpha(1 leads to 3) linkages, but not with any other dextrans, glycogen, and pullulan. Inhibition tests with various sugars show that the combining site of myeloma protein CAL20 TEPC1035 is most complementary to panose, a trisaccharide DGlc alpha(1 leads to 6)DGlc alpha(1 leads to 4)DGlc. Panose was 3.3 times more potent than a tetrasaccharide DGlc alpha(1 leads to 6)DGlc alpha(1 leads to 4)DGlc alpha(1 leads to 4)DGlc and 8, 23, 42, > 42 times more active than maltose, nigerose, isomaltose, and kojibiose, respectively. These findings were paralleled by their binding properties as determined by affinity electrophoresis. The association constants (Ka) of these three dextrans to myeloma protein CAL20 TEPC1035 ranged from 3.8 X 10(3) ml/g to 5.02 X 10(3) ml/g. The association constant of inhibitor (Kia) of panose was 8.19 X 10(3) M-1. Myeloma protein CAL20 TEPC1035 is an antidextran with specificity different from those of other murine myeloma antidextrans and from human antidextrans reported previously and its combining site size is at least as large as a trisaccharide. The binding constant of methyl alpha-D-glucoside (7.2 X 10(2)) was 73% of that of panose and comparable to that of myeloma protein W3129 (9.4 X 10(2)) with a cavity-type site and 600 times lower (1.6 X 10(0)) for QUPC52 with a groove type site, indicating that the terminal nonreducing residue is held in a cavity. Inhibition data with various alpha(1 leads to 4)-linked oligosaccharides also indicate that the internal portions of these inhibitors may react directly with a portion of the combining site. These findings suggest that myeloma antidextran CAL20 TEPC1035 has a partial cavity-type combining site in which the terminal nonreducing dGlc alpha(1 leads to 6) moiety is held in a cavity with the other two sugars forming a groove. However, oligosaccharides with one or more alternating [leads to 3DGlc alpha(1 leads to 6)DGlc alpha(1 leads to 3)DGlcl leads to] units with and without terminal nonreducing DGlc alpha(1 leads to 6) or DGLc alpha(1 leads to 3) side chains remain to be tested to determine whether structures known to be present in the three dextrans which precipitate CAL20 TEPC1035 may not prove to be more active than panose.

Animals↗
PubMed1977

A potential diagnostic method for food allergy: clinical application and immunogenicity evaluation of an elemental diet.

The use of a simple, hypoallergenic elemental diet would appear well suited for diagnosing food allergy. Vivonex was used in 21 patients (5 to 40 years old) suspected of food allergy or those who had failed to respond to the usual management of inhalant allergy. To study immunogenicity, five New Zealand rabbits were immunized with Vivonex, milk, and egg and were evaluated for the production of precipitin and passive cutaneous anaphylactic antibodies, the latter was evaluated in three Hartley guinea pigs. The clinical study was conducted over a 2- to 3-week period with evaluation of symptom and medication scores, physical examination, and hematological and biochemical measurements made before and after the Vivonex trial, which was a minimum of 1 week. No consistent, significant improvement of allergic manifestations were seen while patients received Vivonex. On the other hand, there were no serious side effects noted either clinically or by laboratory measurements, although four patients discontinued the study because of Vivonex palatibility. Vivonex was not immunogenic by either the precipitin reaction or passive cutaneous anaphylactic response. Although Vivonex did not prove helpful in these severe, refractory allergic individuals, we were encouraged by its safety and acceptance in the outpatient setting. Further studies in young allergic children who are more likely to have clear-cut food sensitivity are being planned.

Adolescent↗
PubMed1978

Countercurrent immunoelectrophoresis in the diagnosis of viral infections of the central nervous system.

Countercurrent immunoelectrophoresis was utilized in the study of 621 specimens of cerebrospinal fluid to determine the correlation of detection of viral antigens with the clinical diagnosis of aseptic meningitis and related viral infections. A panel of viral antisera was immunoelectrophoresed against 119 specimens from patients with suspected viral infections of the central nervous system (group I), 32 from patients with bacterial meningitis (group 2), and 470 from patients with no suspected infection of the nervous system (group 3). One or more precipitin bands were detected in 79% of specimens from group 1, 19% from group 2, and 4% from group 3. Paired acute- and convalescent-phase sera from 32 (78%) of 41 patients with precipitin bands detected by countercurrent immunoelectrophoresis demonstrated a fourfold or greater change in complement-fixing antibodies to the detected antigen. With refinements in antisera, countercurrent immunoelectrophoresis may become useful in the rapid laboratory diagnosis of viral infection of the central nervous system.

Acute Disease↗
PubMed1988

Primary cytomegalovirus infection in pregnancy: comparison of antibody responses to virus-encoded proteins between women with and without intrauterine infection.

Using radioimmunoassay followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, we compared serial IgG precipitin antibody responses to cytomegalovirus (CMV) proteins in three groups of 29 pregnant women who had primary CMV infection. Five women had CMV mononucleosis, and four of them infected their fetuses. Twenty-four women had subclinical infection, and eight infected their fetuses. There were no qualitative differences in the precipitin responses against the virus-encoded proteins in three different infected cellular antigens (cytoplasmic, nuclear, and high-speed pellet) between these three groups of women. There was also no qualitative difference in responses whether the infection was clinically apparent or subclinical. Quantitation by densitometer, however, revealed that women with mononucleosis and those with subclinical infection who infected their fetuses had a more intense and prolonged antibody response than did women with subclinical infection who failed to transmit CMV in utero.

Antibodies, Viral↗
PubMed1977

Immunochemical studies on non-precipitating guinea pig antibody produced by administration of an excessive dose of Bacillus subtilis alpha-amylase.

Administration of an excessive dose of Bacillus subtilis alpha-amylase [EC 3.2.1.1, alpha-1,4-glucan 4-glucanohydrolase] (BalphaA) induced the production of non-precipitating (non-ppt) IgG2 antibody in guinea pigs, whereas immunization with a normal dose produced precipitating (ppt) IgG1 and IgG2 antibodies. The non-ppt IgG2 antibody thus produced could be isolated from the coexisting ppt IgG2 antibody by means of the precipitin reaction at maximum precipitation. The non-ppt antibody was incapable of forming a precipitin arc with BalphaA in a conventional agar plate. In the presence of 4% polyethylene glycol (PEG), however, it formed a single arc which fused completely with those of the ppt IgG1 and IgG2 antibodies. The non-ppt antibody could not fix complement, but inhibited BalphaA activity, though with less efficiency than the ppt antibodies. These properties of the non-ppt IgG2 antibody may be due to a low affinity for BalphaA, since both gel filtration and precipitation of soluble antigen-antibody complexes with 20% PEG showed that the antibody was easily dissociable from BalphaA.

Amylases↗
PubMed1982

Antigenic structure of adenylate kinase from porcine skeletal muscle. I. Three immunochemically active peptides obtained by cleavage with cyanogen bromide.

Analysis of the quantitative precipitin reaction of adenylate kinase from porcine skeletal muscle with goat anti-adenylate kinase antiserum indicated that there are at least four antigenic determinants on the enzyme molecule. Porcine adenylate kinase was cleaved with cyanogen bromide, and four peptides were fractionated by ion-exchange chromatographies. Three fragments, CBb (2-56), CBfN (81-125), and CBfC (126-194), inhibited the quantitative precipitin reaction of intact adenylate kinase with goat antiserum. CBb, CBfN, and CBfC also inhibited the binding of 125I-labeled adenylate kinase to the specific antibody purified from goat antiserum. In both inhibition studies, the inhibitory activity of each fragment was extremely high, and reached 70% or more in the latter case. From these results and in view of the presence of the sequence -Glu-Glu-X-X'-Lys (or Arg)-Lys- in each immunochemically active fragment, we suggest that these fragments have similar antigenic determinants which are cross-reactive.

Adenylate Kinase↗
PubMedDate not supplied

Detection of IgG candida antibodies by solid-phase radioimmunoassay and comparison with agar-gel diffusion.

Comparison of solid-phase radioimmunoassay (S-PRIA) and agar-gel diffusion for detecting antibodies to candida organisms was undertaken in patients with systemic candidiasis, candida colonization, noncandida febrile illnesses, and in healthy controls. Radioimmunoassay results were highly reproducible when the same sera were tested on three different days. Sera from all three groups of patients had higher counts per minute (CPM) by S-PRIA then sera from control subjects. Although positive precipitins and high CPM occurred frequently in patients with systemic candidiasis, both precipitins and high CPM were detected in some sera from patients with colonization without system disease.

Adrenal Cortex Hormones↗
PubMed1977

Immunological investigations of Candida albicans in respiratory disease.

1. Tests against rabbit antiserum to Candida albicans showed about 40 protein antigens in cytoplasmic extract compared with two in a commercial test extract, which contained mainly polysaccharide (mannan) antigens. 2. Positive precipitin tests to the mannan preparation were recorded in 40% of 56 asthmatics on systemic corticosteroids and 37% of 56 rhinitis patients not so treated. The titres were slightly higher in the corticosteroid group. There was no increase in precipitins over 0-9 months in asthmatics treated with beclomethasone dipropionate aerosol (BDA) by inhalation, or over 0-12 months in rhinitis patients treated with BDA nasal spray. No positive reactions to the purified protein fraction were recorded. 3. Radioallergosorbent tests for specific IgE antibody showed a higher incidence of positive results with whole C. albicans yeast cells than with purified cytoplasmic protein. There was a higher incidence of positive results to protein in patients on systemic corticosteroids than in the other patients. No change in incidence occurred in asthmatics on BDA. The number of positive results in 56 rhinitis patients increased on BDA from 28 to 37 for yeast cells, and from 7 to 16 for cytoplasmic protein. This was attributed to C. albicans challenge tests. 4. Specific IgA antibodies to yeast cells were detected in 48/56 asthmatics on systemic corticosteroids, and there were no significant changes over 0-9 months on BDA. The corresponding figures for rhinitis patients were 25/56, and again there were no significant changes over 0-12 months on BDA.

Antigens↗
PubMed1982

Precipitating anti-VIII:C antibodies in two patients with haemophilia A.

IgG from the plasmas of two haemophilia A patients with anti-VIII:CAg antibodies (1000 and 200 u/ml) was isolated and labelled with 125I. The specific labelled anti-VIII:CAg IgG was further purified by binding to and elution from immobilized factor VIII/von Willebrand factor (F.VIII/vWF). When studied by immunodiffusion and autoradiography, both antibodies gave a precipitin line with normal plasma, serum, cryoprecipitate, purified F.VIII/vWF and the plasmas of two patients with haemophilia A+. No precipitin line was observed with the plasmas of 11 patients with haemophilia A- or four patients with severe von Willebrand's disease. Levels of VIII:CAg obtained by radioelectroimmunoassay were in agreement with those obtained by immunoradiometric assay. This study demonstrates that, contrary to previous evidence, human anti-VIII:CAg antibodies are precipitating as well as neutralizing when studied by highly sensitive techniques.

Antibodies↗
PubMed1976

Radioallergosorbent test (RAST) studies with insolubilized polysaccharides.

Polysaccharide preparations from Aspergillus fumigatus and Candida albicans when used with some human sera were found to fix complement and precipitate with specific antibodies present in the sera. Precipitins consisted predominantly of IgG with traces of IgM and IgA. Following insolubilization of the fungal polysaccharide preparations with the N-carboxyanhydride of L-leucine, reactivity with serum precipitins was retained and reaction with IgE antibodies was demonstrated using the radioallergosorbent test (RAST). Results showed that for A. fumigatus and C. albicans at least, polysaccharide-specific IgE antibodies probably occur in sera from some hypersensitive individuals. We suggest that chemical methods of the type described which induce polymerization or cross-linking will be useful for detecting IgE antibodies in the RAST. Such an approach could be used with a variety of polysaccharide preparations which cannot be coupled to polysaccharide solid supports by the usual methods.

Antibodies, Fungal↗