Manufacturer's approach in transferring accuracy to multiple field installations.

Manufacturers of instrument systems for in vitro diagnostic use are closely regulated by the Food and Drug Administration and sell their products to clinical laboratories that participate in rigorous quality control and proficiency testing programs. In these circumstances, manufacturers must plan and implement procedures that will ensure acceptable accuracy and precision from all installed systems. This activity starts with the design goals and specifications and continues with manufacturing and quality assurance testing, through various postsale support services. This article outlines some specific instrumentation examples to demonstrate the accomplishment of these objectives.

Electrophoretic techniques in today's clinical laboratory.

The clinical laboratory today is being asked to do more tests faster than ever before. The field of electrophoresis is no different than any other part of the clinical laboratory. With these demands comes the need to get more information in a more timely fashion. Agarose gives the clinical electrophoresis laboratory a tool for resolving proteins in a more meaningful manner. These patterns, when confirmed with specific protein analysis and immunofixation electrophoresis, can lead to earlier diagnosis and treatment. Those who say they have no time to run electrophoresis probably have not run electrophoresis lately and are missing out on a very valuable and cost-effective tool.

Major issues in the organization and implementation of the National Cooperative Gallstone Study (NCGS).

The National Cooperative Gallstone Study (NCGS) was a cooperative, randomized, controlled trial of a drug, chenodiol, for the medical dissolution of gallstones. The design and procedures of the NCGS were complex, having developed as a result of extensive involvement of many experts in the field of gallstone disease and biliary lipids. During the design and implementation of the protocol, many important issues required consideration and resolution. The aim of this article is to review these issues and the deliberations surrounding their resolution and provide personal conclusions and recommendations that may be helpful to other investigators involved in cooperative, controlled trials.

Lactic dehydrogenase activity in human pheochromocytoma.

Lactic dehydrogenase (LDH), a cytosolic enzyme found in neural and endocrine tissue, was measured in serum and tumor tissue of 15 patients with pheochromocytoma, a neuroendocrine neoplasm. Mean serum total LDH activity was higher in patients with pheochromocytoma than in patients with essential hypertension, normotensive control subjects, or those with various other categories of secondary hypertension. The prominent isoenzyme was LDH type 3. Their pheochromocytoma tissue, as well as normal human adrenal tissue, also contained LDH, maximally type 3; the amount of LDH in tumors far exceeded that in normal adrenal glands, suggesting that the tumor tissue is the source of the excessive serum LDH in these patients. While a large percentage of false-negative results (40%) does not render serum LDH activity a reasonable screening test for pheochromocytoma, and even though the true-positive rate is high (100%), we cannot yet recommend that hypertensive patients with high serum LDH activity undergo investigation for this tumor.

Evaluation of a monoclonal antibody-based immunoradiometric assay for prostatic acid phosphatase.

This report evaluates a new immunoradiometric assay for prostatic acid phosphatase in serum, based on a dual monoclonal antibody reaction system (Hybritech-TANDEM). A solidphase antibody binds the acid phosphatase molecule and a second monoclonal antibody to a different antigenic site serves as the 125I-radiolabel. The method was tested on 67 patients with various stages of prostatic carcinoma and 134 patients without the disease. It also was compared with a conventional polyclonal radioimmunoassay (NEN) and an enzymatic activity method (duPont aca). The upper limit for the TANDEM assay on nondiseased male patients was found to be 2.0 microgram/L. Based on this upper limit of normal, the diagnostic sensitivity of the method for all cases of prostatic carcinoma was 60%. We could not distinguish the enzyme released in abnormal amounts due to benign prostatic hypertrophy and certain nonprostatic malignant diseases from that of prostatic carcinoma. The diagnostic specificity was calculated at 95%. For the clinically undetectable Stage 1 disease, sensitivity was 44% (four abnormal values out of nine cases). The TANDEM procedure is simple to use and reproducible.

Measurement of intracellular amino acids in cultured fibroblasts.

Tissue culture is a valuable tool for the study of cellular amino acid metabolism. However, accurate measurement of intracellular amino acids is difficult. Cells must be washed to remove the culture medium which is rich in amino acids, a procedure which may result in the loss of intracellular amino acids. Collection of the cells of washing and amino acid measurement is further complicated by the adherence of cultured fibroblasts to the culture vessel surface. An uncomplicated, reliable method for the determination of the amino acid content of cultured human diploid fibroblasts is described. The intracellular levels of several amino acids determined by this method are markedly higher than previously reported levels.

Hepatotoxicity of bile acids in rabbits: ursodeoxycholic acid is less toxic than chenodeoxycholic acid.

The hepatotoxic effects of cholelitholytic bile acids, ursodeoxycholic and chenodeoxycholic acids, were compared with each other and with those of lithocholic acid, a known hepatotoxic bile acid, in the rabbit. Male New Zealand white rabbits were fed regular laboratory chow containing ursodeoxycholic, chenodeoxycholic, or lithocholic acid at a concentration of 0.5 per cent (w/w) for 14 days. The control group was fed the chow without added bile acids. The mortality rate was highest (six of 12) in the lithocholate group, intermediate (two of eight) in the chenodeoxycholate group, and lowest (none of six) in the ursodeoxycholate group. Light microscopy of the liver revealed fibrosis, inflammation, and bile duct proliferation in the portal regions in the three experimental groups; however, the lesions in the lithocholate and chenodeoxycholate groups were more severe and often associated with periportal extension of fibrosis and focal necrosis of the parenchyma. In addition, electron microscopy revealed distortion of bile canaliculi, conspicuous bundles of intermediate-sized filaments, expansion of pericanalicular cytoplasmic matrix due to apparent accumulation of microfilaments, prominence of lysosomes, and fragmentation of cisternae of the rough endoplasmic reticulum. These ultrastructural changes were less marked and often absent in the ursodeoxycholate group. The serum L-alanine aminotransferase activity increased 5- to 6-fold in the lithocholate and chenodeoxycholate groups, whereas it remained less than 2-fold of the control level in the ursodeoxycholate group on day 14. The serum lithocholate concentration was markedly elevated to comparable levels in all three groups, whereas ursodeoxycholate was highly increased in the ursodeoxycholate group but undetectable in the other groups at the time of sacrifice. It is concluded that (1) although the oral administration of three bile acids induces hepatic injuries in the rabbit, ursodeoxycholate causes less severe injury than do the other two, (2) the advantage of ursodeoxycholate versus chenodeoxycholate is probably relative rather than absolute, (3) lithocholate formed through metabolic conversion from ursodeoxycholate may be responsible for the most part for hepatotoxicity, and (4) it is possible that the concurrent presence of ursodeoxycholate may mitigate lithocholate's hepatotoxicity.

A candidate reference method for uric acid in serum. I. Optimization and evaluation.

We describe our optimization and evaluation of a candidate Reference Method for uric acid in serum. Reaction parameters were optimized for a manual, enzymic method for uric acid in which highly purified microbial uricase is used to quantitate uric acid by a differential ultraviolet procedure. We evaluated the method in terms of freedom from interferences, analytical recovery, precision, and comparison with five other uric acid methods. We conclude that (a) the candidate uric acid Reference Method exhibits the least interference; (b) all six methods exhibit satisfactory analytical recoveries and precision; and (c) results by all six methods agree well. As a result of this evaluation study, the manual ultraviolet uricase method for uric acid, with Tris as buffer, was chosen as the candidate Reference Method for uric acid.

A candidate reference method for uric acid in serum. II. Interlaboratory testing.

We describe the interlaboratory testing of a candidate Reference Method (Part I) for uric acid in serum. The method is based on the ultrasound spectrophotometric quantitation of uric acid before and after incubation with uricase. A comprehensive investigation involving 12 laboratories was organized to document the transferability, intra- and interlaboratory precision, and general reliability of the candidate Reference Method. The interlaboratory CV with this method was about 2 to 6% for uric acid concentrations ranging from 0.12 to 0.60 mmol/L. The results detailed here demonstrate that the method can be successfully duplicated among different laboratories.

Rapid development of large numbers of alpha-fetoprotein-containing "oval" cells in the liver of rats fed N-2-fluorenylacetamide in a choline-devoid diet.

Fischer rats, fed 0.05% w/w N-2-fluorenylacetamide in a choline-devoid diet for 2 weeks, develop a massive infiltration of the liver by small "oval" cells. This occurs rapidly one week after feeding the diet for two weeks. All rats fed choline-devoid diet die within 5 weeks, with massive oval cell infiltration of the liver. Although similar changes occur in rats fed N-2 fluorenylacetamide in a choline-supplemented diet, their degree is much less. In rats fed a choline-devoid diet without N-2-fluorenylacetamide, proliferation of hepatocytes, but not of oval cells, is observed. Because the carcinogen-enhancing effects of choline-devoid diets seem to exceed those of partial hepatectomy, such diets may work by causing changes distinct from those induced by partial hepatectomy. Many oval cells contain alpha-fetoprotein, and the rapid oval cell increase is associated with an exponential increase in serum alpha-fetoprotein concentration. These observations suggest that a cellular change, not an alteration of gene expression in parenchymal cells, is the primary cause of hyper-alphafetoproteinemia during the course of chemical carcinogenesis in rats.

Amino acid analysis of elastin hydrolysates using a lithium citrate gradient: quantification of elastin from whole lung.

A method to quantitate the elastin content in lungs is described. Desmosine and isodesmosine, the cross-linking amino acids unique to elastin, were measured directly from acid hydrolysates of hamster and human lungs by amino acid analysis. A Durrum D-500 (Dionex) amino acid analyzer, with a modified lithium citrate buffer gradient was used. Results compared favorably with those obtained by gravimetric and microenzymatic techniques, and the method described could be used for the quantification of elastin in biopsy specimens

Incorrect least-squares regression coefficients in method-comparison analysis.

The least-squares method is frequently used to calculate the slope and intercept of the best line through a set of data points. However, least-squares regression slopes and intercepts may be incorrect if the underlying assumptions of the least-squares model are not met. Two factors in particular that may result in incorrect least-squares regression coefficients are: (a) imprecision in the measurement of the independent (x-axis) variable and (b) inclusion of outliers in the data analysis. We compared the methods of Deming, Mandel, and Bartlett in estimating the known slope of a regression line when the independent variable is measured with imprecision, and found the method of Deming to be the most useful. Significant error in the least-squares slope estimation occurs when the ratio of the standard deviation of measurement of a single x value to the standard deviation of the x-data set exceeds 0.2. Errors in the least-squares coefficients attributable to outliers can be avoided by eliminating data points whose vertical distance from the regression line exceed four times the standard error the estimate.