Determination Of Human Serum Albumin Research Articles

Synthesis, characterization and luminescent properties of Eu III and Tb III fluorescent chelates used as label in medical immunoassays

DTPA-pAS, EDTA-pAS, DTPA-pAB and EDTA-pAB are synthesized and confirmed by elemental analysis, IR spectra, 1H NMR and mass spectra. Fluorescent properties are investigated by mixing the four chelants with Eu IIIor Tb III chloride in aqueous solution. The fluorescence intensity varies with the chelants, pH value and concentration. Only DTPA-pAS and EDTA-pAS enhance the Tb III luminescence efficiently and the fluorescence intensity varied slightly at pH 6.0–12.0. The fluorescence of Tb III-DTPA-pAS can be detected even at a concentration of 10 −7 mol l −1. Tb III-DTPA-pAS has been used as a label in fluoroimmunoassay for the determination of human serum albumin (HSA).

Highly sensitive spectrophotometric determination of human serum albumin with 3',4',5',6'-tetrachlorogallein-molybdenum(VI) complex.

A highly sensitive spectrophotometric determination of human serum albumin (HSA) with 3',4',5',6'-tetrachlorogallein (T.Cl.Gall)-Mo(VI) complex in a Triton X-100 + polyvinyl alcohol micellar medium is proposed. This method can be used to determine up to ca. 150 micrograms/10 ml of HSA from the optical absorbance at 640 nm, and is superior in sensitivity to the other extremely sensitive spectrophotometric methods. The great sensitivity of this method results from the use of third-derivative spectrophotometry. The binding parameters of T.Cl.Gall-Mo(VI) complex to HSA are n = 77.3 and K = 1.05 x 10(4) M-1 as determined from dual double-reciprocal plots. It is suggested that the colored complex in this system may be the association complex between [HSA]m+ and [MoVI(T.Cl.Gall)2]n- involving hydrophobic interaction between HSA and T.Cl.Gall. The proposed method should also be useful for the detection and determination of some peptides (e.g. low molecular weight peptides containing basic amino acids), as well as proteins.

Fluorimetric determination of human serum albumin with eriochrome cyanine R.

A sensitive spectofluorimetric method has been developed for the determination of human serum albumin (HSA). The complex of Eriochrome Cyanine R (ECR) with HSA fluoresces at 423 nm (with excitation at 308 nm). The limit of determination is 0.5 µg ml–1 and the relative standard deviation is less than 2%. The composition of the fluorescent complex is 1 : 4 (HSA : ECR).

A sensitive spectrophotometric method for the determination of human serum albumin with chrome-azurol S aluminium chelate.

A simple and highly sensitive method for the determination of serum albumin based on the formation of a ternary complex from chrome-azurol S aluminium chelate was developed. A nonionic detergent, polyethylene glycol-p-nonylphenyl ether n=10, acted as a stabilizer of the ternary complex in the present system. The formation of the ternary complex caused a large increase in absorbance at 630 nm, and the calibration curve was linear between 10 and 200 μg of albumin in the final colored solution (8.0 ml). The sensitivity of the present method was about eight times that of a bromcresol green control method. Heparin and bilirubin did not interfere with the determination.

Immobilized antibody-based flow type enzyme immunosensor for determination of human serum albumin

A flow-type enzyme immunosensor was prepared for the electrochemical determination of human serum albumin (HSA). The immunosensor was constructed from the immobilized antibody (anti-HSA IgG) reactor and an oxygen electrode. The immunochemical reaction of catalase-labelled antibody with HSA was completed with 30 min. After the immunochemical reaction, hydrogen peroxide solution was injected into the system and a peak current was obtained within 2 min. A linear relationship was observed between the current increase and the logarithm of HSA concentration in the range 10 −8-10 −6 g ml −1. The minimum measurable concentration was 10 −8 g ml −1. The current increase was reproducible with 10% of the relative errors when a sample solution containing 10 −7 g ml −1 of HSA was used. The minimum measurable concentration increased to 10 −9 g ml −1 when hydrogen peroxide was recycled for 5 min in the reactor system. The immobilized antibody reactor could be reused. HSA in human serum was determined by the system proposed.

A catalytic immuno-reactor for the amperometric determination of human serum albumin

A catalytic immuno-reactor for the determination of human serum albumin (HSA) was constructed by using immobilized antibody and an amperometric detector. A sandwich assay with hemin-labeled antibody to catalyze the decomposition of hydrogen peroxide was used, the catalytic activity of the hemin-antibody conjugate being determined by measuring the decrease in hydrogen peroxide concentration. The reaction of hemin-labeled antibody with antigen was complete within 30 min and the current decrease was correlated with the HSA concentration. The relative standard deviation was about 9% at an HSA concentration of 1 mg ml −1.

Solid-phase luminescent catalyst immunoassay for human serum albumin with hemin as labeling catalyst

A solid-phase luminescent catalyst immunoassay is described for the determination of human serum albumin (HSA) in solution; hemin is used as a label which catalytically amplifies the sensitivity. The method is essentially a non-radioactive and non-enzymatic sandwich immunoassay. Anti-HSA antibody is covalently bound to a transparent plate, which then undergoes the immunochemical reaction with HSA in the test solution, and with the fixed amount of hemin-labeled anti-HSA antibody. After the two-step immunoreaction, the immunochemically-adsorbed hemin-antibody conjugate is quantified by means of the luminescence produced in a solution containing luminol and hydrogen peroxide. The luminescence intensity is correlated with the amount of HSA. The limit of detection for HSA is 1 ng ml -1.

Ultrathinfilm immunoelectrophoresis.

Immunoelectrophoresis (Laurell-rocket) in ultrathinfilm gels has been applied to the determination of human serum albumin in a model study. Agarose gels of 0.24 mm thickness were used and optimum electrophoretic conditions were determined. Precision was assessed and comparison with conventional immunoelectrophoresis and a chemical method was made. Though precision was poor (CV 4-6%) improvements in dispensing small volumes (0.2 microliter) should change this. Good correlation between methods (correlation coefficient 0.89-0.97) was obtained. The advantages of the technique are speed of analysis and low reagent costs.

An approach for the quantitation of serum albumin utilizing the principle of protein-dye binding

An amidoschwartz 10B slope method is described for the determination of human serum albumin. The method is based on the principle that amidoschwartz binds differently to albumin and globulin fractions, thereby giving standard curves with different slopes. A linear relationship between the slope and the albumin-to-total protein ratio was obtained, which allows the calculation of the albumin concentration. The method was found to be highly reproducible (coefficient of variance = 3.6%) and gave a reference value for albumin and an A G ratio in good agreement with values previously reported.

Thermometric enzyme linked immunosorbent assay: TELISA

A new method, thermometric enzyme linked immunosorbent assay (TELISA), for the assay of endogenous and exogenous compounds in biological fluids is described. It is based on the previously described enzyme linked immunosorbent assay technique, ELISA, but utilizes enzymic heat formation which is measured in an enzyme thermistor unit. In the model system studied determination of human serum albumin down to a concentration of 10 −10 M (5 ng/ml) was achieved, with both normal and catalase labelled human serum albumin competing for the binding sites on the immunosorbent, which was rabbit antihuman serum albumin immobilized onto Sepharose CL-4B.

The determination of human serum albumin by its specific binding ofthe anionicdye, 2-(4′-hydroxybenzeneazo)-benzoic acid

A method of improved sensitivity and reliability is described for the direct determination of human serum albumin using the anionic dye, 2-(4′-hydroxybenzeneazo)-benzoic acid. Some important factors of the albumin-dye reaction are discussed. A pH 6.2 phosphate buffer system has several advantages over a pH 5 acetate buffer system; the quality of the dye should be established; temperature control is necessary; and careful attention should be given to the choice of a human albumin product as a reference standard, for which only human mercaptalbumm or human albumin-Fraction V can be recommended.