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Biomedical subjects

R Malvano

Publications and source records attributed to R Malvano.

At least 37 records · Page 2Linked to original sources
PubMed1979

Creatine kinase forms in human skeletal and cardiac muscle.

The creatine kinase (CK) activity in human skeletal and cardiac muscle submitted to QAE-Sephadex chromatography was found to distribute into three peaks (m and h I, II, III fractions). Characterization according to electrophoretic behaviour, approximate molecular weight, thermal stability and immunological properties unambiguously demonstrated the coincidence of fractions I and III with reference MM-CK and MB-CK isoenzymes, while both cardiac and muscular forms II proved to escape any assignment within the dimeric (M, B) model. A higher molecular weight than for reference CK and the absence of interaction with BB-antiserum resulted for both forms II, which on the contrary were found to diverge as for reactivity to MM-antiserum and stability characteristics. Quantitation of the CK forms, attempted on a limited number of samples, confirmed the expected relative levels of MM-CK and BB-CK and gave evidence for the presence of hII- or mII-CK amounting up to 30% of the total activity in cardiac and skeletal muscle, respectively. The results of the study both confirm a complexity greater than supposed for the CK system and point to the inadequacy of several of the commonly used analytical approaches to derive correct information.

Chromatography, Ion Exchange↗
PubMed1979

Radioimmunoassay of lactate dehydrogenase, H forms.

Antisera to H4-lactate dehydrogenase (LDH) were elicited in rabbits, against both human (h) and porcine (p) isoenzymes. 125I-labelled H4-LDH was prepared by electrolytic iodination. A simple and fast procedure (1-h incubation for clinical assays) was set up by using polyethylene glycol for the bound-free separation. The results obtained in the antiserum characterization indicated that the heterologous homotetramer, M4 was completely discriminated in the porcine system, while a weak cross-reaction with human antisera resulted. In both cases, for the hybrid forms, a cross-reactivity level related to the stoichiometric contents of the H-subunit in the tetramers was observed. The H4-LDH from other species was found to be much more effectively disinguished in the porcine than in the human system. The assay for human LDH was further validated in terms of analytical suitability and clinical response. For healthy subjects the mean concentration was 0.46+/-19 micrograms/ml (mean+/-SD). Patients with acute myocardial infarction had levels ranging from 1.2 to 5.9 micrograms/ml.

Humans↗
PubMed1979

Radioimmunoassay of unconjugated and total serum estetrol using a 125I-iodinated tracer.

A radioimmunossay (RIA) for the measurement of both unconjugated and total serum estetrol has been developed, using an antiserum to an E4-3-conjugate and a 125I-radioiodinated E4 tracer. Assay of dried ethyl ether extracts was used for the determination of unconjugated E4, while a direct measurement of unextracted hydrolyzed serum in the presence of 0.3% 8-anilino-1-naphthalene sulphonic acid (ANS) proved adequate for total E4. Assay reliability was evaluated and the procedure standardized through a series of tests aimed at assessing accuracy, sensitivity and precision. No steroidal interference was found to practically affect the assay (0.3% estriol cross-reactivity), nor were solvent and sample blanks observed in the case of unconjugated E4. For total E4 assay, the sample blank effects were acceptably overcome by using hydrolyzed male serum and 0.3% ANS, as a standard diluent. An interassay variability amounting to approximately 10 and 6% resulted for unconjugated E4 and total E4 RIA, respectively. A number of serum samples (285 for unconjugated E4, 147 for total E4) randomly collected throughout normal pregnancy were assayed. The unconjugated E4 levels at 15th week and at term were 62.7 +/- 22.6 and 766.5 +/- 208.2 (SD) pg/ml, respectively. Total E4 was about 6--7 times higher than the levels of free E4 and increased 7 times from the 15th week to term.

Circadian Rhythm↗
PubMed1977

Direct radioimmunoassay of estradiol in serum extracts.

The feasibility of direct radioimmunoassay of unconjugated estradiol in dried serum extracts in the assessment of ovarian function in the regulation of the menstrual cycle has been investigated. Using an antiserum to an estradiol-6-conjugate, assay reliability was evaluated and the procedure standardized through a series of tests aimed at assessing accuracy, sensitivity, and precision. These included multiple titration with competing steroids, the definition of blank effects from solvent and sample, the assay both of samples to which known amounts of estradiol had been added and of serially diluted samples, and clinical validation using as a reference samples related to well-defined physiological situations. The variability of replicate estimates and the repeatability of the calibration curve were evaluated to obtain information on assay precision and sensitivity. The analytical performances proved to be perfectly adequate for the clinical purposes for which the assay was intended, in terms both of reliability of results and of methodologic practicability.

Animals↗
PubMed1976

Solid-phase radioimmunoassay of plasma triiodothyronine.

A direct radioimmunoassay method for circulating triiodothyronine was set up, using an insolubilized (cellulose-coupled) antiserum and 8-anilino-1-naphthalene-sulfonic acid (ANS) as a protein inhibitor. Particular attention was paid to the evaluation of changes in the physico-chemical properties of the antibody sites under the experimental conditions adopted: no substantial decrease of immunoreactivity was found to be associated with the insolubilization process, while complex effects following the use of protein inhibitors were observed. The standardized procedure was defined in terms of analytical reliability both by a comparison with a reference (charcoal-dextran) method and by evaluation of sensitivity, precision and accuracy.

Anilino Naphthalenesulfonates↗
PubMed1976

Direct radioimmunoassay of plasma cortisol.

The simplification of the measurement of circulating cortisol by direct radioimmunoassay of plasma samples sets the problem of inhibiting the carrier proteins competing with antibodies. This was accomplished by exploiting the much higher effectiveness of pH and temperature variations on steroid binding to carrier proteins than to antibody sites. A solid-phase system was set up, using antisera to cortisol-21-BSA conjucates coupled to CNBr-activated cellulose. The standardized procedure consisted of an incubation at pH 3.5 and room temperature, directly assaying 10 mul of plasma. A methodological and clinical validation of the measurement was carried out through a series of tests aimed at assessing the reliability of results (assay of steroid-deprived plasma, recovery test and serial dilution of samples, comparison between different antisera and with different methods including extraction, responsiveness to well-established physiological situations). The results obtained are reported and the validity of the method discussed in terms of more general applicability to steroid assay.

Adrenocorticotropic Hormone↗
PubMed1976

Methodological simplifications in radioimmunoassay of urinary aldosterone.

Simplification of radioimmunoassay procedures of urinary aldosterone-18-glucuronide was attempted, taking into consideration the aspects implied by the hydrolysis of urine and the assay itself. The procedure standardized for the hydrolysis step (samples diluted with a two-fold volume of 0.2 N HCl and incubated at 30 degrees C for 16-24 h) proved suitable in terms of practicability and accuracy. Aldosterone antisera, raised in the rabbit against an aldosterone-3-bovine albumin conjugate, were selected according to their specificity towards competing steroids. Depending on the characteristics of the antisera used, an assay of extracts, or even direct measurements of hydrolyzed urines excluding any extraction, were found to yield reliable results. In the case of a high-quality antiserum, evidence for the adequacy of assay on non-hydrolyzed urine extracts for the measurement of the excretion of unconjugated aldosterone was provided by some preliminary data. The results of the experiments, directed at the methodological and clinical validation of the simplified procedures, are reported and discussed in this paper.

Aldosterone↗
PubMed1975

Measurement by radioimmunoassay of unconjugated estriol in pregnancy serum.

Antisera to estriol 6--onjugates were tested for suitability in the direct radioimmunoassay of unconjugated estriol in extracts of pregnancy serum. Assessment of specificity through titration with competing steroids allowed a selection to be made within the group of antisera. The measurement was then standardized by checking the absence of analytical blank (solvent and sample blank) and the extent of binding variability associated with the use of charcoal-dextran as a separating agent. Validation was made by means of the usual recovery and dilution tests, and by a cross-comparison of analytical data obtained with different antisera; reproducibility of calibration curve and within-and between-assay variability was evaluated under routine conditions. The validity of the clinical information was assessed by assaying 202 samples randomly collected throughout normal pregnancy from the 16th week to term: both trend and levels of unconjugated estriol concentration were found to be in good agreement with the literature data.

Animals↗
PubMed1975

Antiserum characteristics and assay quality.

On the basis of the experience derived from the setting up and standardization of different radioimmunoassay systems, this paper attempts to point out the role played by the characteristics of the antiserum in the optimization of the assay. In particular the antiserum heterogeneity, the rates of formation and dissociation of immunecomplexes, and the nature of the antigen-antibody bonds are taken into consideration as regards some implications on the extent of blank, the effect of separation procedures and the adequacy of preincubation techniques.

Antibody Specificity↗
PubMed1975

Results of a comparative study on insulin radioimmunoassay in 36 Italian laboratories.

An interlaboratory study in which the insulin contents of five plasma samples were estimated in 36 italian laboratories was coordinated by the Istituto Superiore di Sanità (National Institute of Health) and the Consiglio Nazionale delle Ricerche (National Research Council). A rather large between-laboratory variability resulted, though the ranking of samples according to their insulin concentrations was practically the same. A significant dependence of estimates on the method used was established. The analysis of data, aimed at defining the possible reasons of the assay variability, is reported and discussed.

Insulin↗
PubMed1971

Preparation and immunoreactive properties of monoiodinated angiotensin labelled at high specific activity.

Monoiodinated angiotensin II at a specific activity in the order of 1 C/mg can be prepared with high yields by controlling the pH of iodination and can be purified on G-25 Sephadex. The fractions from the head to the centre of the single peak of radioactivity obtained by gel filtration of the iodination mixture contain pure monoiodinated angiotensin. The monoiodinated derivative shows an immunoreactivity very close to that of the native hormone; in contrast, the diiodinated derivative has a low immunoreactivity and its use as a tracer results in a loss of sensitivity in the radioimmunoassay of angiotensin II. It is suggested that the use of the monoiodinated derivative in pure form should be recommended whenever a small hormonal polypeptide labelled at high specific activity is to be prepared for radioimmunoassay purposes.

Angiotensin II↗