Biomedical subjects
Publications and source records attributed to B Mattiasson.
An enzyme immunoassay for the detection of benzo[a]pyrene covalently conjugated to macromolecules has been developed. The monoclonal antibody, raised through in vitro immunization reacted with benzo[a]pyrene metabolites bound to DNA, RNA and proteins. The lower detection limit for the assay was 1 pmol for benzo[a]pyrene bound to DNA or RNA, and 5 pmol when bound to protein.
A developmentally regulated carbohydrate-binding protein from the capture organs of Arthrobotrys oligospora, not present on hyphae, was isolated and partially characterized. Surface structures of A. oligospora were radiolabeled with [125I]iodosulfanilic acid. The fungus was homogenized, and the homogenate was passed over an affinity column containing N-acetyl-D-galactosamine immobilized to Sepharose 6B. The bound radiolabeled protein was eluted from the affinity column with a glycine-hydrochloride buffer (pH 3.0), concentrated, and chromatographed on a metal chelate affinity gel containing Ca2+. EDTA was used as an eluant for the radiolabeled protein. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis in combination with autoradiography revealed a molecular weight for the carbohydrate- and cation-binding polypeptide of ca. 20,000.
Partitioning in aqueous 2-phase systems was used to separate free and bound ligand in an immunoassay for beta 2-microglobulin. In order to get efficient separation in the phase system, the antibodies were modified to favour their partition in a different phase from that of antigen. However such modification of antibodies significantly decreased their binding capacity. This was overcome by using antibodies bound to previously modified staphylococci, which had proper partitioning behaviour. Alternatively, antibodies conjugated with biotin could be used in combination with modified avidin. This paper presents a method for the evaluation of data from immunoassays whereby 2-phase systems have been used to separate free and bound antigen.
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A radioimmunoassay for triiodothyronine T3 and thyroxine T4 without any manual separation is described. By the use of an aqueous two-phase system the free hormone was separated from the immunologically bound fraction and lifted out of the counting area of the gamma counter. The coefficient of correlation between results obtained with this method and with a conventional RIA was 0.97 and 0.93 for T3 and T4, respectively.
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A time dependent decrease of sample counts was observed in an adsorption radioimmunoassay with internal sample attenuator counting. The drift caused a bias in the estimate of the amount of antigen in the samples. The size of this deviation was dependent upon the length of time after the calibration curve was made that the samples were measured. This was essentially due to dissociation of the antigen-antibody complexes as the adsorber can act as a second receptor to the antigen. Addition of slowly sedimenting starch microspheres or starch particles inhibited the drift by forming a diffusion barrier on the attenuating pellet.
Partitioning in two-phase systems was used as a separation method in binding analyses and compared with equilibrium dialysis. A phase system was chosen to give partitioning conditions with the reactants in different phases. The interaction between serum albumin and Cibacron Blue F3G-A was used as a model system. The protein partitioned to the bottom phase and the ligand to the top phase in a phase system consisting of poly(ethylene glycol) and dextran. When both albumin and Cibacron Blue were present in the phase system, the degree of binding was observed as a translocation of ligand from the top to the bottom phase. An evaluation method that compensates for the amount of free reactant in the phase with the binding protein is presented.
A simple rapid assay for microorganisms is described. An adsorbent capable of binding microbial cells is deposited in a disposable plastic syringe. A sample to be analyzed is drawn into the syringe and the number of attached microorganisms is calculated from their ability to produce metabolites. The smallest number of yeast cells in a given sample was 50,000. Each analysis takes 2.5 h.
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Enzymic determination of L-ascorbic acid, D-galactose, D-glucose, cellobiose, lactose, and sucrose with an enzyme thermistor is described. The enzyme thermistor is a simple flow-calorimeter designed for routine analysis and measures metabolites by the heat produced in a small column with immobilized enzyme. This instrument may be used for determination of discrete samples as well as for continuous monitoring. As the change of enthalpy produced by the primary reaction is usually large enough to make the measurement sufficiently accurate, simple assay procedures can be designed, an advantage of great importance when the samples are crude and complex.
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Radioimmunoassay of triiodothyronine was performed with a homogeneous incubation system. Bound and free ligand were separated by partitioning in an aqueous two-phase system. The agreement between the results obtained with this technique and conventional heterogeneous radioimmunoassay was very good (r = 0.981).
A new method is described for detection of membrane antigens and antibodies against such structures. By inserting partially purified rat transplantation antigen (RT-1) into iodine or fluorescein-labeled lipid vesicles, a precipitation of the liposome is demonstrated by the use of a specific alloantiserum and a heterologous anti-rat IgG serum. Precipitation of liposomes carrying WF transplantation antigens could be detected at a final dilution of the alloantiserum of 1:3000, which is comparable to that obtained in a 51Cr-release assay. Furthermore, when unpurified membrane proteins from WF splenocytes were incorporated into labeled liposomes, an amount of RT-1 corresponding to 8000 cells could be detected.