Cationic latex particles with surface amino groups were prepared by a multistep batch emulsion polymerization. In the first one, two or three steps, monodisperse cationic latex particles to be used as the seed were synthesized. In the third and fourth steps, the amino-functionalized monomer aminoethylmethacrylate hydrochloride was used to synthesize the final functionalized latex particles. Three different azo initiators 2,2Ј-azobisisobutyramidine dihydrochloride, 2,2Ј-azobisdimethylenisobutyramidine dihydrochloride, and 2,2Ј-azobisisobutyronitrile were used as initiators. Hexadecyltrimethylammonium bromide was the emulsifier. To characterize the final latices, conversions were obtained gravimetrically, and particle size distributions and average particle diameters were determined by transmission electron microscopy and photon correlation spectroscopy. The amount of amino groups was determined by conductimetric titrations. Colloidal aspects were ascertained by measuring the electrophoretic mobilities. Activation of these particles with glutaraldehyde produced an efficient reagent for latex-enhanced immunoassay. The covalent coupling efficiency (protein covalently bound with respect to the total amount of protein adsorbed) was compressed between 50 and 80%. The developed immunoreagent was applied to the measurement of serum ferritin concentration in a new turbidimetric procedure that was compared with a commercial nephelometric method; the results obtained with both methods demonstrated that the two procedures correlated well (r ϭ 0.992).
N-terminal pro-brain natriuretic peptide (NT-proBNP) and BNP measurement could have a significant role in differentiating dyspnea between cardiac or pulmonary origin in the emergency room. The development of new and different commercial assays for these B-type natriuretic peptides offers the possibility of improving and simplifying their measurements but this could be defaulted due to the differences in methodology and the lack of assay standardization. We compared four available methods of measuring NT-proBNP and BNP and evaluated their usefulness in diagnosing the causes of breathlessness in the emergency room. The correlation of BNP with different assays was strong with r>0.98 (P<0.0001). Comparison studies between NT-proBNP and BNP procedures were in good agreement with r>0.87. The area under the receiver-operating characteristic curve (ROC) for BNP or NT-proBNP for detecting any cardiac dysfunction was higher than 0.96 (95% CI). A BNP value of 116 pg/mL measurement with the Access BNP assay (Beckman Coulter Inc., Fullerton, CA), a BNP value of 79 pg/mL with Advia Centaur BNP assay (Bayer Diagnostics, Tarrytown, NY), and an NT-proBNP level of 817 pg/mL in Elecsys NT-proBNP assay (Roche Diagnostic, Mannheim, Germany), showed both high sensitivity (>92%) and high specificity (>93%). We have found that NT-proBNP and BNP present similar diagnostic accuracies for the differential diagnosis of dyspnea.
We have developed a new procedure for turbidimetric measurement of ferritin concentration in human serum, based on latex microparticle agglutination technology. The procedure has been automated using the Falcor 300 analyzer. Carboxilated latex particles (336 nm in diameter) were covalently coupled with immunopurified F(ab')2 fragments of anti-ferritin IgG antibodies. Coated microparticles were automatically mixed with undiluted sample and the resulting absorbance due to agglutination was measured at 550 nm. The procedure generated a calibration curve with a measuring range of 0 to 558 microg/l, showing a day-to-day imprecision lower than 5.7%. The detection limit was 4 microg/l. There were no interferences from bilirubin, hemoglobin or rheumatoid factors. Turbid and lipemic samples caused an important interference which could be avoided by pretreating those samples prior to measurement. A prozone effect was provisionally obtained with ferritin concentrations over 1800 microg/l. The results suggested a hook-like effect due to a rapid microparticle precipitation in the reaction media, that could be avoided by increasing the reaction medium density by adding sucrose to the buffer, up to 150 g/l concentration. This sucrose addition resulted in a displacement of the Heidelberger curve with a prozone phenomenon occuring at concentration higher than 3000 microg/l of ferritin. Results obtained with the present procedure correlated well with those obtained by a nephelometric procedure and with those obtained by an RIA. We conclude that this latex turbidimetric immunochemical procedure is simple, rapid, has a good analytical and operational performance on the Falcor 300 analyzer and is well suited for the measurement of ferritin concentration in human serum.
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