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Biomedical subjects

C G Magnusson

Publications and source records attributed to C G Magnusson.

At least 55 records · Page 3Linked to original sources
PubMed1986

Raised serum IgG4 levels in patients with atopy and filariasis: application of an automated particle-counting immunoassay using monoclonal antibody.

We show here an automated (50 samples/h) assay for serum IgG4 having a throughput time of 40 min per sample and a sensitivity of 10 micrograms/ml. The assay procedure is based on the inhibition by sample of the agglutination reaction between monoclonal anti-IgG4 antibodies and latex particles to which IgG4 myeloma protein has been coupled. Assay reliability was ascertained by testing for linearity, analytical recovery (96.4%), interassay precision (less than or equal to 8%), specificity and correlation between the results obtained with monoclonal and polyclonal anti-IgG4 antibodies (n = 84; rs = 0.97). Application of the assay to sera from various groups of patients indicated significantly (p less than 0.00005) higher geometrical means (Gx) in patients suffering from atopy (n = 87; Gx = 617 micrograms/ml), atopic dermatitis (n = 28; Gx = 1,043 micrograms/ml), filariasis with Onchocerca volvulus (n = 48; Gx = 1,681 micrograms/ml) and Brugia malayi (n = 20; Gx = 1,078 micrograms/ml) as compared to nonatopic subjects (n = 103; Gx = 302 micrograms/ml) and randomized paired maternal/cord sera (n = 41; Gx = 276 and 296 micrograms/ml, respectively). IgG4 in the paired maternal/cord sera correlated (r = 0.98; p less than 0.00005). There was no significant influence of age or sex on the IgG4 levels either among the nonatopics or the atopics even though low IgG4 (less than or equal to 30 micrograms/ml) was more common among women. The results suggest that IgG4 and IgE responses are somehow closely related in atopic and parasite-infested patients at the physiological, pathogenic or genetic level.

Adolescent↗
PubMed1986

Histamine release from human leukocytes by anti-IgE antibodies: influence of multiple or single epitope recognition.

Upon comparing several polyclonal anti-IgE antisera (specific for the Fc epsilon fragment) for their ability to release histamine, we observed a marked heterogeneity of response for the same antiserum in different cell donors as well as for different antisera in the same cell donor. Additional studies with monoclonal antibodies directed against various epitopes of the D1 and D2 regions revealed several possible explanations: broad specificity of the polyclonal antisera, lack of availability of some epitopes on the cell-bound IgE (which were available on soluble IgE), microheterogeneity among IgE molecules from different patients, and failure of some donors' cells to respond to anti-IgE antibodies. We conclude that selection of appropriate antisera is an important consideration in comparing IgE-mediated histamine release among individuals.

Antibodies, Anti-Idiotypic↗
PubMed1986

Monoclonal antibodies against human IgE. Identification of a determinant restricted to IgE of the lambda light-chain type.

The specificity of five monoclonal anti-IgE antibodies (Mabs) was studied in direct latex agglutination and agglutination-inhibition experiments by particle-counting immunoassay. Twenty IgE myeloma proteins and several purified D epsilon O-, D epsilon 2-containing pepsin and papain fragments of IgE-DES(kappa) were used in the evaluation. The results demonstrate two Mabs with isotypic specificity for two distinct epitopes of the Fc epsilon-fragment within the D epsilon 1- and D epsilon 2-determinants. One Mab recognized only the immunizing IgE protein and was directed against determinants on the Fd epsilon-fragment probably related to the idiotype. Anti-Em(1) allotypic Mabs recognized all 20 IgE myeloma proteins including two of Japanese origin and the Em(1)-allotype was confined to D epsilon-determinants. Interestingly, one Mab (ALE) reacted with all 8 IgE myeloma proteins of the lambda light-chain type but none out of 12 bearing kappa chains. ALE seems therefore to recognize a new marker on IgE besides the known idiotypic, allotypic and isotypic ones. These results illustrate that a critical specificity control of Mabs is always warranted. Moreover, one should be aware of possible interference in IgE assays from the kind of determinants recognized by ALE whenever intact IgE myeloma proteins are used to raise polyclonal antisera, to get immunosorbent-purified anti-IgE antibodies or when used as tracers and standards.

Antibodies, Monoclonal↗
PubMed1985

Inhibition of idiotype-anti-idiotype reaction in particle counting immunoassay as a tentative assay of IgE.

Rabbit antibodies were raised against a purified mouse monoclonal IgG1 antibody having specificity for the D epsilon 2 determinant of human IgE. After appropriate absorption this antiserum was anti-idiotype specific and was used to set up a particle counting immunoassay for human IgE. Latex particles coated with F(ab')2 fragments of the anti-idiotypic antibodies were agglutinated by the monoclonal anti-IgE antibody (20 ng/ml). This reaction was inhibited up to 100% by human IgE and the assay took 30 min to perform with a sensitivity of 40 IU/ml. The coefficient of correlation with a routine IgE assay was r = 0.96, and the mean analytical recovery tested on 10 sera was 95.8%.

Antibodies, Anti-Idiotypic↗
PubMed1985

Immunoglobulin E in aqueous humour.

The level of immunoglobulin E (IgE) was determined in the aqueous humour (AH) and serum by particle counting immunoassay (limit of sensitivity: 0.2 IU ml-1). This level in the AH was below 0.75 IU ml-1 in case of senile cataract (n = 22) and ranged from this assumed normal value to 550 IU ml-1 in case of uveitis (n = 37), Fuchs' cyclitis (n = 11), intraocular tumour (n = 7) and perforating ocular injury (n = 6). To know whether the IgE increase was related to a possible local production the AH/serum concentration ratio for IgE was calculated and divided by that for albumin. Values higher than 0.60 were considered as suggestive of a local synthesis and were observed in 31 of the 37 samples examined.

Aqueous Humor↗
PubMed1985

Immunoglobulin E assayed after pepsin digestion by an automated and highly sensitive particle counting immunoassay: application to human cord blood.

We describe in this article a fully automated, universal assay for serum IgE after pepsin digestion of the sample and subsequent assay of the released Fc" epsilon fragment by particle counting immunoassay (PACIA). The sensitivity and the range of the assay were easily modulated by changing the concentration of dextran in the reaction medium. In the application to cord serum, the sensitivity reached 0.1 IU/ml in 30 min incubation time and with a throughput of 50 analyses per hour. Within-day and between-day coefficients of variation did not exceed 7.6% for IgE levels covering a wide range of the standard curve. Dependable accuracy was demonstrated by linearity tests, analytical recoveries (89% to 112%), and correlation with PRIST on 48 samples from children ages 1 to 14 yr (y = 0.97x + 5.92; r = 0.987). Minor discrepancies between the two methods were attributed to a slight serum effect in PRIST. PACIA applied to 348 cord serum samples demonstrated a statistically significant influence of sex and race on the cord IgE level. In European neonates boys had significantly (p = 0.019) higher geometrical mean cord IgE levels (n = 142; 0.46 IU/ml; range less than 0.10 to 30 IU/ml) than girls (n = 146; 0.33 IU/ml; range less than 0.10 to 8.0 IU/ml), which was attributed to a predominance of boys (chi 2 = 4.29; p less than 0.05) having more often elevated cord IgE (less than 1.20 IU/ml) than girls. Neonates of African-Asian origin had significantly (p less than 0.00005) higher cord IgE levels (n = 60; 1.05 IU/ml; range less than 0.10 to 125.0 IU/ml) than European neonates (n = 288; 0.39 IU/ml; range less than 0.10 to 30.0 IU/ml).

Animals↗
PubMed1985

Clinical and immunological aspects of a case of monoclonal hyper-IgE. Isolation of IgE-protein, estimation of basophil cell bound IgE and histamine release.

A case of monoclonal IgE type lambda with IgE levels about 1 mg/ml has been followed for 6 years. Except for a slight asthma no signs of malignancies, parasitic infestations or other known diseases compatible with hyper-IgE have been found. By the combination of fractional ammonium sulphate precipitation, gel filtration, chromatofocusing, and subtraction affinity chromatography the IgE protein was isolated in an immunochemically pure and homogeneous form. Immunofluorescence of bone marrow cells showed about 1% IgE plasma cells. The amount of basophil bound IgE was 42 ng/10(6) cells, and histamine release from basophils challenged with anti-IgE was not different from that in atopic control persons, indicating a within-allergic-patients normal amount of IgE receptors. The protein-A reactivity was 0.4% equivalent to well-known IgE myeloma proteins. No antigen specificity of the IgE protein was found. Only a few cases of asymptomatic hyper-IgE are known, and it cannot be ruled out that this represents a premyeloma condition, since a similar case terminated in a malignant lymphoma.

Aged↗
PubMed1985

A reappraisal of IgE levels in various human secretions by particle counting immunoassay combined with pepsin digestion.

We have assayed IgE in various human secretions by particle-counting immunoassay (PACIA). The introduction of a pepsin digestion step of the sample prior to the assay not only eliminates possible interfering factors but also releases from IgE the protease-resistant Fc"epsilon fragment. Since this fragment could represent the residual form of IgE in certain secretions rich in proteases it should be a suitable calibrator for the assay of IgE in secretions. The concentrations of IgE that we found in most fluids (urine, saliva, tears, milk, aqueous humour, cerebrospinal fluid, broncho-alveolar washings, intestinal fluids and faecal extracts) were up to 100 times lower than those reported by others who have used the radioimmunosorbent test (RIST). We suspect that the high values found with RIST are related to the susceptibility of this test to various interfering factors and not to a local production of IgE in most secretions of normal individuals.

Body Fluids↗
PubMed1984

Typing of subclasses and light chains of human monoclonal immunoglobulins by particle counting immunoassay (PACIA).

The subclasses of monoclonal IgGs and IgAs were identified by particle-counting immunoassay. The principle of the test is the inhibition of the agglutinating activity of either specific antisera or monoclonal antibodies (for IgA only) on latex particles coated with a monoclonal IgG or IgA of known subclass. The feasibility of assay of polyclonal Ig subclasses was demonstrated. However, the anti-IgG2 antiserum cross-reacted with an allotype (nG4m(b)) of IgG4. The possibility of typing monoclonal Igs for light chains by the same technique was also demonstrated. Results are obtained in 30 min, and the method requires only small amounts of purified immunoglobulins (Igs) and antisera or monoclonal antibodies.

Agglutination Tests↗
PubMed1984

IgE in the cerebrospinal fluid.

The level of IgE in the cerebrospinal fluid (CSF) was determined by particle counting immunoassay. With a limit of sensitivity of 0.2 IU/ml, this immunoglobulin was detected neither in CSF of non-neurological patients (n = 27) nor of patients with sciatica (n = 13). IgE was present in samples from some patients with either multiple sclerosis (MS) or various infections of the central nervous system. In these cases, an IgE index [CSF IgE/serum IgE: CSF albumin/serum albumin] was calculated as 0.29 (SD 0.12). This value is not abnormal as the mean IgG and monomeric IgA indices are 0.45 and 0.34, respectively. Therefore, the IgE detected in most of the CSF samples was not locally produced. However, most patients with tuberculous meningitis had clearly an increased IgE index suggesting a local biosynthesis, but we failed to detect any IgE antibody activity against purified protein derivatives.

Antibody Specificity↗
PubMed1984

A three-layer immunoradiometric assay for determination of IgG subclass antibodies in human sera ("IgG subclass RAST"). Validation of the subclass specificity, and establishment of equipotency.

We report the development of a three-layer immunoradiometric assay (TIRA) for measurement of IgG antibodies of all four subclasses in human sera. The first layer consists of diluted human serum, the second layer is monoclonal mouse antibodies to human IgG subclasses, and the third layer is 125I-labelled rabbit anti-mouse IgG. Monoclonal anti-IgG1, anti-IgG3 and anti-IgG4 reacted only with their complementary IgG subclass, whereas the anti-IgG2 showed slight cross-reactivity to immunoglobins of other subclasses and classes and to light chain proteins. The observed cross-reactivity was found to be without importance, when the TIRA was applied to measurement of IgG subclass antibodies. Equipotency was established by use of appropriate dilutions of the monoclonal antibodies, and the assay was calibrated by use of human reference serum. The TIRA therefore permits reliable inter-individual and intra-individual comparisons of the IgG antibody response in all four subclasses. Non-specific binding obtained with pooled normal human serum was below 0.33%. Inter-assay coefficient of variation was between 18 and 27%. The TIRA was applied to measurement of IgG subclass antibodies to timothy grass pollen in sera from grass pollen allergics undergoing immunotherapy.

Animals↗
PubMed1983

An automated particle counting immunoassay (PACIA) for determination of blocking antibodies against timothy grass pollen in sera from desensitized allergics.

An automated particle counting immunoassay (PACIA) for measurement of blocking antibodies (antigen neutralizing capacity) against timothy grass pollen extract in sera from desensitized allergics is described. Latex particles coated with F(ab')2-anti-timothy are agglutinated by timothy. Serum containing anti-timothy antibodies inhibits the agglutination. Non-agglutinated particles are counted in a modified AutoCounter. Nineteen of 20 sera from timothy allergics who had undergone immunotherapy with purified timothy extract for 30 weeks, showed significant agglutination-inhibition. None of 42 normal human sera gave significant inhibition. The inhibiting antibody could be removed by absorption with protein A and was thus of non-IgE nature, i.e. blocking antibody. The results obtained correlated statistically significantly with those found with a double-antibody method (rS = 0.62, n = 20, t = 3.35, P less than 0.01) and with the cumulated dosage of timothy allergen extract administered to the individual patient (rS = 0.56, n = 20, t = 2.87, P less than 0.02). Between-assay coefficient of variation was from 6.4% to 18.3%. The capacity is 40 samples per hour. The method has also been applied to measurement of blocking antibodies to honey bee and wasp venom.

Animals↗
PubMed1983

The use of particle counting immunoassay (PACIA) for the titration of specific antibodies. Application to sera from honeybee venom-desensitized patients.

Particle counting immunoassay (PACIA) is shown to be a suitable nonradioisotopic method for the titration of anti-phospholipase A2 antibodies in the sera from honeybee venom-desensitized patients. The sensitivity of the method is equal to the RAST-IgG with a correlation coefficient between the two methods of r = 0.94. Within-day and between-day precisions were 2.5 and 9.2%, respectively. The main advantages of PACIA are: (i) the use of easily prepared non-radioactive reagents having a shelf-life of at least 1 year at -20 degrees C: (ii) partial automation; (iii) calibration with antisera of animal origin, and (iv) the possibility of allergen standardization of honeybee venom extracts.

Agglutination↗
PubMed1983

Activation of the classical pathway of human complement by a human monoclonal IgE, IgE(DES).

Activation of the classical pathway of human complement by monoclonal IgE from patient DES was demonstrated by using IgE(DES) coupled to latex particles. This material depleted human serum of C1 and C4 hemolytic activities. In addition, C3bi was deposited in a calcium-dependent way onto the insolubilized IgE as shown by the agglutination of latex by conglutinin. The alternative pathway was also activated. These anticomplementary activities were dose and time dependent. Moreover, we confirmed that another monoclonal IgE, IgE(PS), activated the alternative pathway exclusively. Particular attention was paid to exclude contamination by other immunoglobulins or C-reactive protein, generation of artifacts due to the chemical coupling, and the presence of proteolytic enzymes in the IgE(DES) preparation. Moreover, evidence is also presented against the involvement of IgG or IgM anti-IgE autoantibodies that could activate the classical pathway after their binding to insolubilized IgE(DES). Although one cannot exclude the possibility that IgE(DES) or IgE(PS) are abnormal proteins, these findings suggest the existence of an isotypic or allotypic variation of IgE.

Animals↗
PubMed1982

Particle counting immunoassay of immunoglobulin E antibodies after their elution from allergosorbents by pepsin: an alternative to the radioallergosorbent test.

The absorption of specific antibodies on allergen-coated cellulose discs, followed by pepsin digestion to release the IgE Fc" fragment, which is then assayed by particle counting immunoassay (PACIA), is a new practical and reliable method to quantitate IgE antibodies and to standardize allergens. The coefficient of variation of repeated assays does not exceed 12.8% and the correlation coefficient with RAST was r = 0.96. Some discrepancies could be explained by differences in the antiserum specificities and the presence of anti-IgE autoantibodies. The determination of IgE antibodies by PACIA offers the following advantages: (1) no radioisotopes; (2) stability of reagents, since antibody-coated latex particles and the standards keep their activity for more than 1 yr; (3) low cost of reagents due to ease of preparation --PACIA does not require purified antibodies to measure IgE but only F(ab')2 fragments of the IgG fraction of the anti-IgE antiserum to coat latex; (4) avoidance of nonspecific absorption of the labeled anti-IgE on the solid phase; (5) prevention by elution of the inaccuracies observed in RAST at very high titers of IgE antibodies; (6) expression of the results in international units per milliliter to calculate the ratio of specific IgE Ab vs total IgE, with a high ratio suggesting that the patient is sensitive to a few allergens; (7) standardization of allergenic extracts; and (8) the particular specificity of the anti-IgE antiserum directed against a heat- and protease-resistant fragment of IgE.

Allergens↗
PubMed1982

Particle counting immunoassay (PACIA) of ferritin.

Particle Counting ImmunoAssay (PACIA) has been adapted to the determination of serum ferritin. Polystyrene particles (0.8 micrometers) were coated with anti-ferritin antibodies and the concentration of ferritin determined by the agglutinating activity of this protein. The agglutination was measured by the reduction of the number of non-agglutinated particles counted in an optical blood cell counter. Non-specific agglutination and inhibition of agglutination were prevented in 99% of clinical samples by the use of F(ab')2 fragments of the antiserum IgG to coat the particles and by using slightly dissociating conditions (pH 9.2, ammonium thiocyanate, EDTA). The system was automated with a sampling rate of 50/h and an incubation time of 25 min. The standard curve ranged from 1 to 100 micrograms/l; recoveries were between 93.6 to 100.2%; the correlation coefficients of PACIA with radioimmunoassays calculated respectively on 99 and 91 samples were 0.974 and 0.984; maximal within- and between assay CV were 11.2% and 7.7%.

Agglutination↗
PubMed1981

An automated particle-counting immunoassay (PACIA) for serum IgE.

Total IgE was determined in 107 sera using a novel, automated, non-isotopic immunoassay called PACIA (particle-counting immunoassay) based on agglutination of anti-IgE coated latex particles by IgE. The IgE values ranged from 10 to 50,000 iu/ml and were compared with results obtained by a conventional radioimmunoassay (RIA) which used a fast double antibody separation technique--the coefficient of correlation was 0.985 and the regression line y = 0.82 x +130.00. PACIA had several advantages over the RIA technique: using a sampling rate of 50/hr, results were obtained in 35 min compared to 16-20 hr, no labelled IgE was required and the separation step, which relied on measuring the number of non-agglutinated particles by an optical cell counter, was fully automated. The threshold of sensitivity was 10 iu/ml and the maximal coefficient of variation for between assay precision was 12.9%.

Humans↗