Laser Nephelometry Research Articles

The measurement of amniotic fluid immunoglobulins by laser nephelometry.

Using laser nephelometry IgG, IgA, and IgM levels in amniotic fluid samples from normal pregnancies were measured between 15 and 24 weeks and between 37 and 41 weeks gestation. Not enough samples between 25 and 36 were available for statistical analysis. IgG levels increased until the 20th and 24th weeks of gestation and declined significantly at term. However amniotic fluid IgM and IgA levels rose at term. Five samples of amniotic fluid associated with a fetus of abnormal karyotype were examined. In one case of Down's syndrome excessive IgA and IgM levels were found. In amniotic fluid obtained after intrauterine fetal death at term, very high levels of all three immunoglobulins were found.

Serum protease inhibitors in acute viral hepatitis

Serum levels of alpha 1-antitrypsin (alpha 1-AT) and alpha 2-macroglobulin (alpha 2-M) and, as controls, alpha 1-acid glycoprotein (alpha 1-AG) and haptoglobin were evaluated by means of laser nephelometry in 17 patients with acute viral hepatitis (AVH) type A, 16 with AVH-B, 12 with AVH-NANB and 8 with fulminant hepatitis B. On admission, alpha 1-AT levels were elevated in one third of AVH-A and AVH-B cases, but subsequently declined; alpha 2-M levels were elevated in about 40% of AVH-B patients during the 2nd, 3rd and 4th week after admission. No significant correlation was found between elevated levels of protease inhibitors and aminotransferase values or drug addiction and delta coinfection. alpha 1-acid glycoprotein and haptoglobin levels were always normal or low. Protease inhibitors did not show any elevation in fulminant hepatitis, while changes were found only in a few patients with AVH-NANB. Thus, no clearcut pattern of changes in protease inhibitors has been found in association with each type of hepatitis, although alpha 1-AT and alpha 2-M elevations are mainly found in AVH-B.

Measurement of α-Thiol Proteinase Inhibitors in Hmman Semm by Laser Nephelometry

Measurement of α-Thiol Proteinase Inhibitors in Hmman Semm by Laser Nephelometry

Ascitic fluid and serum C-reactive protein concentrations in patients with and without peritonitis.

Seventy-five paired ascitic fluid and serum specimens were tested for C-reactive protein (CRP) concentrations with the use of laser nephelometry. There was no clear separation of the ascitic fluid or serum values of the 19 paired specimens obtained from patients with peritonitis from the 37 paired sterile portal-hypertension-related samples or from the 19 paired miscellaneous specimens. The ascitic fluid CRP concentrations of patients with sterile portal-hypertension-related ascites were not significantly different from those of infected specimens. However, the serum CRP values were significantly higher in patients with peritonitis than in patients with sterile portal-hypertension-related ascites. Ascitic fluid CRP does not appear to be a useful indicator of ascitic fluid infection.

Food antibodies, intestinal permeability and HLA status in IgA deficient blood donors identified by a new rapid screening test.

Circulating food antibodies, intestinal permeability and HLA status were studied in twelve blood donors previously identified as being selectively IgA deficient from screening 10,000 donations by means of a latex agglutination inhibition test, and confirmed by laser nephelometry. Assessment of intestinal function also included clinical history and standard laboratory tests. The donors proved to be healthy with no evidence of autoimmune disease or allergy. Nine donors (75%) had precipitins to food antigens. None had increased intestinal permeability as measured by the cellobiose/mannitol absorption test. HLA-A, B and DR antigen distribution was normal. IgG sub-class distribution was normal with the exception of an IgG2 level slightly below normal in one donor (0.9 g/l). An unexpected finding was a return of IgA levels to normal in four donors (33%) within 6 months.

Reference intervals for eight plasma proteins in preterm and term newborns by laser nephelometry.

Reference intervals for eight plasma proteins in preterm and term newborns by laser nephelometry.

Permeability and secondary membrane formation of a high flux polysulfone hemofilter

It has been assumed that the molecular weight (MW) cut-off of a newly fabricated polysulfone capillary dialyzer (F60, Fresenius, FRG) is similar to that of the human glomerulus. We recently tested the device in vivo and found this not to be so, based on the device's ability to eliminate substances of a MW of 10,000 to 60,000 daltons. One of the reasons for this discrepancy was found to be the influence of secondary membrane formation on solute permeability. Endogenous marker substances of a defined MW (beta 2-microglobulin, myoglobin, RBP, alpha 1-microglobulin, acid alpha 1-glycoprotein, alpha 1-antitrypsin, prealbumin, and albumin were measured by laser nephelometry or radioimmune assay; sieving coefficients (SC) and protein eliminations were calculated for each low MW protein.

An enzyme-linked immunoassay for direct measurement of the gelatin-binding capacity of human plasma fibronectin

A new solid-phase enzyme immunoassay measuring the gelatin-binding capacity of plasma fibronectin has been developed. This assay is based on the direct and high-affinity interaction between fibronectin and gelatin coated to polyvinyl chloride plates. The amount of fibronectin bound to gelatin is then measured by sequential incubation with a specific rabbit anti-human fibronectin antiserum, with horseradish peroxidase-conjugated goat anti-rabbit IgG antibodies and with substrate. The final degradation of the substrate is read at 492–650 nm in an ELISA processor. The assay allows the accurate detection of fibronectin concentrations ranging from 1 to 20 μg/ml, is inhibited by the addition of gelatin to plasma, is highly reproducible (interplate CV < 10%), requires 100 μl of plasma only and has been fully automated. Significant linear correlations were noted between total antigenic fibronectin (measured by laser nephelometry) and fibronectin gelatin-binding capacity in plasma from 310 blood donors. Both parameters were higher in men than in women and significantly increased according to age. Dissociation between immunoreactive fibronectin and fibronectin gelatin-binding capacity was observed in two polytraumatized patients. This enzyme immunoassay therefore provides a new method to investigate functional alterations of the gelatin-binding domain of fibronectin in various pathological conditions.

Changes in tear protein composition in Graves' ophthalmopathy

The tear protein composition of normal volunteers and that of patients suffering from hyperthyroidism and various degrees of infiltrative endocrine ophthalmopathy were studied by polyacrylamide-gel electrophoresis, radial immunodiffusion and laser nephelometry. Increased level of IgG, serum albumin and transferrin was demonstrated in hyperthyroidism and severe infiltrative ophthalmopathy. Elevated IgA, IgM and immune complex (IC) levels were detected in cases with mild and severe infiltrative ophthalmopathy, which showed good correlation with the severity of the disease. A decrease in the level of tear specific proteins showed the dysfunction of the lacrimal glands in long-standing cases of Graves' ophthalmopathy. Elevated immunoglobulin levels in cases with irritation are partly the result of increased vascular permeability. High IgA, IgM and 1C levels observed without permeability changes in mild ophthalmopathy suggest the local production of immunoglobulins in infiltrative ophthalmopathy and their possible role in the development of the eye manifestations of Graves' disease. The use of tear protein determinations in the diagnosis and in the evaluation of different therapeutic procedures is discussed.

Antithrombin III measurement in serum and plasma by laser nephelometry may lead to controversial results

Antithrombin III measurement in serum and plasma by laser nephelometry may lead to controversial results

Elevated ascitic fluid fibronectin concentration: A non-specific finding

Ascitic fluid fibronectin concentration was measured in 111 specimens by laser nephelometry. Sterile, portal hypertension-related fluid fibronectin concentration (24 +/- 14 micrograms/ml) was significantly lower than the concentration in infected, portal hypertension-related ascites (49 +/- 44 micrograms/ml, P less than 0.001), peritoneal carcinomatosis (123 +/- 45 micrograms/ml, P less than 0.001), massive liver metastases-related ascites (55 +/- 21 micrograms/ml, P less than 0.001), as well as in ascites of other types (94 +/- 42 micrograms/ml, P less than 0.001). The percentage of samples with fibronectin concentration, greater than 75 micrograms/ml, was 0% for sterile, portal hypertension-related ascites, 28% for infected, portal hypertension-related ascites, 89% for peritoneal carcinomatosis, 20% for massive liver metastases-related ascites, and 72% for ascites of other types. Ascitic fluid fibronectin concentration correlated in a linear fashion with ascitic fluid total protein (r = 0.81, P less than 0.001). Fibronectin concentration in ascites appears to be elevated under a variety of conditions and does not appear to be a specific marker for cancer.

Interaction of the alpha-mannosidase from Canavalia ensiformis with the lectin from the same plant, concanavalin A.

The specific interaction between the lectin concanavalin A and the alpha-mannosidase from the Leguminosa Canavalia ensiformis was studied by means of laser nephelometry and affinity chromatography. Both proteins react optimally within a certain stoichiometrical range. Interaction is restricted to a narrow pH interval (around pH 5) and to low ionic strengths (less than 10mM NaCl). Neither the sugar-binding site of the lectin nor the catalytic and the hydrophobic sites of the enzyme participate in the interaction. The conformation of the enzyme at pH 5 which favours the interaction can be arrested by immobilization. After this, the enzyme is able to bind the lectin even at pH 8 where no interaction takes place between the dissolved proteins.

The Clinical Evaluation of Plasma Fibronectin as a Marker for Nutritional Depletion and Repletion and as a Measure of Nitrogen Balance

Plasma fibronectin has been suggested as a possible marker for nutritional repletion or depletion. This study was undertaken to evaluate the usefulness of plasma fibronectin in patients who received intense nutritional support. Twenty-seven patients referred to our Nutritional Support Services were followed for 3 to 5 wk; 22 received parenteral hyperalimentation alone, two received enteral alone, and three received a combination of both. Plasma fibronectin, serum albumin, serum transferrin, total lymphocyte counts, and 24-hr urine nitrogen balance studies were performed weekly; anthropometric measurements were performed every other week. Plasma fibronectin concentration, measured by laser nephelometry, showed a significant rise (p less than 0.005) in all patients after 1 wk of nutritional therapy; however, there was no significant difference among the subsequent weeks. Plasma fibronectin did not correlate with nitrogen balance studies, serum albumin, or total lymphocyte counts. A correlation between serum transferrin and plasma fibronectin was found not to be clinically useful. Thus, plasma fibronectin is sensitive to nutritional repletion after 1 wk of therapy, but is not useful thereafter. The relationship among nutritional status, immunologic function, plasma fibronectin, and other serum proteins are discussed.

LASER NEPHELOMETER AND LATEX DETERMINATION OF CRP SERUM LEVELS IN A PEDIATRIC POPULATION

CRP is an acute phase reactant which rises rapidly during inflammation and infection and whose values can be usefully utilised in the precocious diagnosis of sepsis in infants and children. CRP serum levels are usually determined using two different techniques: laser nephelometry (LN) and latex agglutination test (LA). The former is the most sensitive and specific method but long and expensive, while the latter is a simpler, cheaper and rapid test. The aim of our study was to value if LA had enough sensitivity to be used as a general screening test, even in the wards, and to use LN only in positive latex patients where it is important a serial quantitative determination of CRP. We studied 262 patients between 2 months and 12 years of age, admitted to the Dpt. of Pediatrics, University of Padua, with a suspicion of infection. CRP levels were determined by means of Behring LN module I, type B and by Rapi-tex CRP by Behring Institute. In our laboratory normal value of CRP concentration in newborn serum by LN have been found to be < 1.0 mg%. Accordingly we consider, in agreement with other authors, that value to be normal also for infants and children. In our study LA showed a sensitivity of 67%, a specificity of 93% and a predictive value of 74%. Even if we consider, in agreement with some others authors, < 1.5 mg% the normal LN value of CRP, LA shows a sensitivity of 73% with a specificity of 94% and a predictive value of 76%. We conclude that LA is not as sensitive to be used as screening technique and that actually only LN method is useful for CRP determination.

A simple method for rheumatoid factor quantitation by laser nephelometry

A simple method for rheumatoid factor quantitation by laser nephelometry

Discrepancy of Serum IgA Levels Determined by Single Radial Immunodiffusion and Laser Nephelometry in Patients with IgA Nephropathy

The levels of IgA in sera were quantitated by single radial immunodiffusion (SRID) and laser nephelometer (LN) to evaluate a precise amount of serum IgA in patients with IgA nephropathy, other glomerular diseases and healthy adults. Thirty patients with IgA nephropathy, twenty patients with other glomerular diseases and eighteen healthy adults were examined. It was shown that the levels of IgA in sera measured by LN were significantly higher than those in sera measured by SRID in patients with IgA nephropathy associated with increased levels of serum IgA. This suggests that the levels of serum IgA measured by SRID in patients with IgA nephropathy were affected by the physicochemical characteristics of circulating IgA in patients with IgA nephropathy. It is postulated that an increase of serum IgA in patients with IgA nephropathy may be mainly due to an increase of polymers rather than a monomer of IgA.

The importance of antibody affinity in the performance of immunoassays for antibody

(I) Introduction Over the last 25 years or so, a wide range of new antibody immunoassay techniques has been developed to add to the historically accepted methods such as precipitation, agglutination and complement fixation. These newer techniques involve the use of electrophoresis, fluorescence, radioactivity, laser nephelometry, solid-phase immunoadsorbents and enzyme conjugates. In general, all tests for antibody-antigen interaction exploit the fact that following combination with antigen antibodies will agglutinate cells or particles coated with antigen, form precipitates with the antigen which can be visualized in gel or solution or otherwise quantified, or can potentiate measurable biological reactions. Not surprisingly, there is a wide range of sensitivity of such tests which depends in part on the particular property of the antibody which is being exploited. Thus tests which depend solely on the primary binding of antibody to antigen (e.g., equilibrium dialysis, fluorescence quenching, radioimmunoassays and enzyme-linked immunoabsorbent assays) are more sensitive than those requiting secondary manifestations of binding (e.g., precipitation, agglutination and complement fixation) (Minden et al., 1968). However, it is becoming increasingly clear that the affinity of an antibody for the corresponding antigen markedly influences its biological activity. Thus to characterize an antiserum solely in terms of its antibody content or titre is less th~n adequate and therefore a knowledge of the affinity of the antibody is of crucial importance (Steward and Steensgaard, 1983). It is also apparent that the affinity of the antibody being measured is itself very likely to influence the efficiency of the assay employed for its quantification. In this review, the importance of antibody affinity in the performance of a number of immunoassays is discussed. In particular, emphasis is placed upon the influence of antibody affinity on the enzyme-linked immunosorbent assays (ELISA) which are enjoying considerable international popularity.

Effects of penicillamine on serum immunoglobulins and immune complex-reactive material in primary biliary cirrhosis

Although penicillamine is used in the treatment of primary biliary cirrhosis, its mechanism of action in this disease is unknown. As an immunologic action had been attributed to the drug, we investigated whether penicillamine might alter serum immunoglobulin levels or immune complex-reactive material in patients with primary biliary cirrhosis. Immunoglobulin levels and immune complex reactivity were measured and clinical tests were performed in 53 consecutive patients entering a double-blind randomized trial of 750 mg vs. 250 mg of penicillamine. Measurement of immune complex reactivity was determined by laser nephelometry, 125I-C1q binding, and Raji cell assays. Immune complex reactivity was detected by at least one assay in 75% of patients tested before treatment. Sixty-two percent were positive in the C1q assay, 28% in the Raji cell assay, and 39% by nephelometry. After therapy with either dose, we found no change in immune complex-reactive material by any assay. Concentrations of immunoglobulins G and M fell (p < 0.05) after 12 mo of therapy. Concentrations of immunoglobulin A decreased (p < 0.05) only in the high-dose group. Correlation was not consistent between results of immune complex assays and clinical liver tests. Although immunoglobulin levels fell during penicillamine therapy, no decrease in immune complex-reactive material was detected. The effect of penicillamine in primary biliary cirrhosis is not mediated through alteration of immune complex-reactive material.

Specificity of an antibody to a subunit of high-molecular-weight storage protein from wheat seed and its reaction with other cereal storage proteins (prolamins)

An antiserum to subunit 2 from the high-molecular-weight (HMW) subunits of the glutenin fraction of Triticum aestivum cv. Highbury was shown to react with related subunits from other cultivars of wheat. The reaction was measured quantitatively by laser nephelometry in polyethylene glycol phosphate-buffered saline after dissolving the HMW fraction in 0.1 M acetic acid; urea used to dissolve the HMW prolamins inhibited the reaction, in some cases at the low concentration of 0.06 M. A study of the comparative reactions of other cereal prolamins was made. 'D' hordein, the homologous HMW protein of barley, showed less reaction, which was more inhibited by urea than the wheat subunits. Some ω-gliadins from the wheat cultivars Chinese Spring and Cheyenne reacted more strongly than the injected fraction and there was less inhibition by urea. A-, β- and γ3 of wheat also reacted with the antiserum while a secalin of rye of Mr 40000 gave a weak reaction.

Fibronectin plasma levels after cadaver kidney transplantation.

The fibronectin plasma levels of 17 patients undergoing cadaver kidney transplantation were determined serially in the postoperative course using laser nephelometry. While 9 patients retained their grafts (group A), the grafts of 8 patients had to be removed (group B), mainly due to rejection, 3 patients had severe infections. In both groups a significant drop of the plasma fibronectin occurred after surgery. There were no significant differences between the two groups in the mean fibronectin levels. After the initial drop the group A patients exhibited rising values leading to a stable level. Progressively declining fibronectin plasma concentrations were found in 2 of 3 severely infected patients. Fluctuating values were found in 3 group B patients without a clear correlation to the rejection crises or the kidney function. The data suggest that the fibronectin plasma level does not seem to be a prognostic marker for graft rejection. But it might be useful in the important and often difficult differentiation between rejection and infection.