bluuebunny/crossref_metadata_2025 huggingface dataset:
Use of dna amplification methods for clinical diagnosis in autoimmune diseases
Authors: Fox Robert I. , Dotan Iris , Fei Hong Ming , Compton Teresa , Hamer Merlin , Saito Ichiro
Published: March 2007

Autoimmune disease is generally felt to result from the interaction of genetic and environmental factors. In recent years, significant advances have been made in using recombinant DNA methods to analyze specific genetic factors and infectious agents. However, new techniques are needed that are more rapid, inexpensive, and suitable for small tissue biopsies obtained early in the course of disease. New methods of DNA amplificaiton based on polymerase chain reaction (PCR) and Qβ‐replicase (QβR) have recently been reported. These methods are briefly reviewed, and their potntial applications to patients with autoimmune disease are presented. Several types of applications can be considered, including detection of: a) specific HLA‐D alleles in order to predict prognosis and better utilize existing medications; b) bacterial, fungal, and spirochete infections in joint aspirates or synovial biopsies; c) human immunodeficiency virus (HIV) and other viruses (e.g., EBV, CMV) that may be associated with immune dysregulation in certain patients; and d) neoplastic transformation in blood or tissues by determining monoclonal gene rearrangements, karyotypic alterations or oncogene activation.

It is likely that routine clinical laboratories will soon begin implementing DNA amplification methods in order to screen blood products for infectious agents (especially HIV and hepatitis B virus). Because these techniques will be readily available, rheumatologists/clinical immunologists should begin developing strategies that will allow them to use these methods in a cost‐effective manner for diagnosis and monitoring treatment.
Immune response and epitope mapping of a candidate hiv‐1 p17 vaccine hgp30
Authors: Boucher Charles A. B. , Krone Willy J. A. , Goudsmit Jaap , Meloen Rob H. , Naylor Paul H. , Goldstein Allan L. , Sun Daisy K. , Sarin Prem S.
Published: March 2007

A thirty amino acid synthetic peptide (HGP30) representing the conserved region of HIV‐1 p17 induced high titer antibodies to the native pl7 in rabbits. This immune sera neutralized HIV‐1 replication in cell culture and one of the high titer antisera also inhibited CD4‐dependent cell fusion. Pepscan analysis with overlapping nonapeptides derived from the sequence of HIV‐1 pl7 identified the sequence (KE) ALDKIEE (EQ) as the major antibody binding site. Sera of 9% of AIDS patients (7/76) and 18% of HIV‐1 seropositive healthy homosexuals (40/223) were positive for HGP30 antibodies. Decline in HIV‐1 pl7 antibodies has been shown to be related to disease progression in both children and adults, suggesting that HIV‐1 pl7 antibodies may be protective. Hence, a synthetic HIV‐1 pl7 peptide, representing the immunodominant epitope, could be useful as a candidate vaccine for immunization of HIV‐1 seronegative or seropositive healthy homosexuals.
Functional interactions between interleukin‐4, interleukin‐2, and tumor necrosis factor‐al for lymphokine‐activated killer cell generation
Authors: Blay Jean Yves , Branellec Didier , Robinet Eric , Gay Francoise , Chouaib Salem
Published: March 2007

The purpose of the present study was to explore the interaction between interleukin‐2 (IL‐2), interleukin‐4 (IL‐4), and tumor necrosis factor‐a (TNF) on the differentiation of human large granular lymphocytes (LGL) into lymphokine‐activated killer cells (LAK). The data show that recombinant human IL‐4 (100‐1,000 U/ml) was able to induce the differentiation of human LGL into LAK effectors. The levels of the IL‐4‐induced cytotoxicity are significantly lower than those observed after stimulation of LGL by optimal doses of IL‐2. This LAK activity generation by IL‐4 was not associated with LGL proliferation. When TNF was added in LGL culture in the presence of sub‐optimal concentrations of IL‐4, the lytic capacity of the activated killer cells was significantly enhanced, suggesting an apparent synergy between these two factors. Most interestingly, our data indicate that exogenous TNF can partially overcome the known inhibitory effect of IL‐4 on IL‐Pinduced LGL differentiation into LAK effectors. These findings suggest a role for TNF in the process of LAK induction.
Specific enzyme immunoassay for a‐fetoprotein from hepatocellular carcinoma
Authors: Kohno Takeyuki , Ishikawa Eiji , Kitamura Tooru , Tsuda Kazunori
Published: March 2007

α‐Fetoprotein in sera from healthy subjects and patients with liver cirrhosis and hepatocellular carcinoma was fractionated into three peaks by affinity chromatography on a column of Lens culinaris agglutinin‐Sepharose 48. One peak (the first peak) was found in all the serum samples, but the other two peaks (the second and third peaks) were found only in patients with hepatocellular carcinoma. For α‐fetoprotein in the second and third peaks, a specific enzyme immunoassay was developed. Lens culinaris agglutinin‐coated polystyrene balls were incubated with α‐fetoprotein and, after washing, with affinity‐purified anti‐a‐fetoprotein Fab'‐β‐D‐galactosidase conjugate. β‐D‐galactosidase activity bound to the polystyrene balls was assayed by fluorsimetry. The maximal volume of serum that could be used without interference was 0.2 m̈l, and the minimal detectable serum concentrations of α‐fetoprotein in the second and third peaks were 3.5 mg/liter and 0.1 mg/liter, respectively. The maximal serum concentration of α‐fetoprotein in the first peak that did not affect the detection limits of a‐fetoprotein in the second and third peaks was 10 mg/liter. It was possible to confirm the presence of a‐fetoprotein in the second and third peaks by a significant difference between bound β‐D‐galactosidase activities in the absence and presence of α‐methyl‐D‐mannoside or D‐glucose. This assay may be useful for diagnosis of hepatocellular carcinoma, although further improvements remain to be made.
A new quantitative nitroblue tetrazolium reduction assay based on kinetic colorimetry
Authors: Virella Gabriel , Thompson Tab , Haskill‐Strowd Rebecca
Published: March 2007

A new method for the quantitative assay of nitroblue tetrazolium (NBT) in which the reduction is measured by kinetic colorimetric analysis is reported. The assay is conducted along standard conditions as far as neutrophil isolation and stimulation, except that the test is performed on microtiter plates and the change of color corresponding to NBT reduction is monitored on a kinetic enzyme immunoassay (EIA) reader for 25 min at 490 nm. The results are expressed as mOD/min/in. The influence of several parameters on the results of the assay was studied, including cell concentration, the nature and concentration of the stimulus, and the freshness of the reagents. Cell concentrations of 5 × 106 and 1 × 107/ml were found to be optimal, and IgG‐coated immunobeads, at a concentration of 1 mg/ml, were found to be the ideal stimuli. NBT reduction for nine normal volunteers studied at 5 × 106 cells/ml ranged from 1.80 to 7.30 mOD/min (mean ± SD = 3.66 ± 1.69). NBT reduction values at 1 × 107 cells/ml in six normal individuals ranged from 2.59 to 7.41 (4.73 ± 1.89). In contrast, NBT reduction in a child with clinical symptoms suggestive of chronic granulomatous disease was 0.31 mOD/min. This method is considerably simpler than any alternative method for the performance of quantitative NBT assays.
Antibodies to the metacestode of taenia solium in the saliva from patients with neurocysticercosis
Authors: Acosta E.
Published: March 2007

IgG antibodies to antigens of Taenia solium metacestodes were detected in saliva samples from patients with intracerebral cysticercosis by means of an enzyme‐linked immunosorbent assay (ELISA). When compared for IgG antibody activity, saliva samples from patients with various nonparasitic neurological disorders and from clinically healthy individuals yielded significantly lower (p < 0.001) absorbance values than saliva samples from patients with neurocysticercosis. However, no differences were observed in IgA anti–T. sodium activity between patients with neurocysticercosis and controls. These results indicate that the detection of anticysticercus IgG antibodies in saliva by means of ELISA may be of value in the diagnosis of neurocysticercosis. Moreover, collection of saliva provides a noninvasive sampling method for immuno‐epidemiological surveys on this disease.
Elevated levels of circulating immune complexes in human immunodeficiency virus infection
Authors: Reddy Mohan M. , Grieco Michael H.
Published: March 2007

Circulating immune complexes were quantitated in asymptomatic intravenous drug abusers (IVDA), male homosexuals, and patients with lymphadenopathy or acquired immunodeficiency syndrome (AIDS) by two different methods. Circulating immune complexes were elevated in HIV‐seronegative IVDAs and male homosexuals, probably reflecting viral infections such as cytomegalovirus (CMV). Circulating immune complexes were also elevated in HIV‐seropositive IVDAs and male homosexuals and in patients with lymphadenopathy or AIDS, reflecting infections such as CMV and HIV.
Impairment of jacalin binding to serum IgA in IgA nephropathy
Authors: Andre P. M. , Le Pogamp P. , Chevet D.
Published: March 2007

A test was set up to analyze the direct binding of serum IgA to the lectin jacalin. Under the testing conditions, jacalin bound to both IgA subclasses and reacted similarly with monomeric and polymeric IgA. A jacalin index was defined to quantify serum IgA binding to this lectin. The jacalin index appeared significantly lower in IgA nephropathy than in controls. This may be related to abnormal IgA glycosylation, which could explain, at least in part. the mesangial deposition responsible for the renal disease.
Assay methods for cyclosporine monitoring following liver transplantation
Authors: Haven Mary C. , Sobeski Linda M. , Earl Robert A. , Markin Rodney S.
Published: March 2007

This article reviews therapeutic drug monitoring for cyclosporine in liver transplantation. Brief descriptions of various immunoassay methods include sample matrix selection, assay reagents, and metabolite cross‐reactivity information. Multiple comparisons of the various methods are outlined. Examples of the method‐dependent relationship between clinical events and changes in cyclosporine concentration are presented. Other potential predictors of liver allograft function are listed.
A dot‐elisa for the detection of igg antibodies to mumps and varicella viruses
Authors: Leonardi Maria S. , Gazzara Domenica , Zummo Sebastiana , Mastroeni Pasquale , Fattal‐German Michèle , Bizzini Bernard
Published: March 2007

An enzyme‐linked immunosorbent assay using nitrocellulose strips (dot‐ELISA) for the routine laboratory detection of IgG antibodies to mumps and varicella viruses is described. The virus antigens are dotted onto nitrocellulose strips, and the dotted strips are incubated with the sera to be tested. The bound antibodies are revealed using enzyme‐labeled antihuman IgG antibodies. Reliable results are obtained when the assay is carried out at 37°C. The reported data indicate that the dot‐ELISA can reliably be used for the detection of IgG antibodies to mumps and varicella viruseis in human sera.
Multiparametric analysis of peripheral blood in the normal pediatric population by flow cytometry
Authors: Wiener Doris , Rowlands David T. , Lowell Nancy , Shah Shirish , Malone John , Lowitt Saul
Published: March 2007

Lymphocyte subset analysis was performed on 114 healthy children and 84 healthy adults. Samples were prepared by a whole blood lysis technique and analyzed by flow cytometry. The percentage and total number of CD3 +, CD4 +, CD8 +, and CD19 + lymphocytes were calculated for each of six age groups. A direct correlation with age was seen in the percentages of CD3 +, CD4 +, and CD8 + lymphocytes. The absolute number and percentage of total lymphocytes, the percentage and absolute number of CD19 + lymphocytes, and the absolute number of CD3 + lymphocytes decreased with age. No significant correlation with age was observed for white blood cells, the absolute number of CD4t and CD8+ lymphocytes, and the CD4 + /CD8+ ratio.
Human, rabbit, bovine, and porcine creatine kinase isoenzymes are glycoproteins
Authors: McBride James H , McBride James H , Rodgerson Denis O , Hilborne Lee H.
Published: March 2007

Human, rabbit, bovine, and porcine creatine kinase (CK) isoenzyme preparations were extensively purified by isoelectric focusing and high‐performance liquid chromatography and tested for the presence of carbohydrate. In this study, all the CK isoenzymes demonstrated positive periodic acid‐Schiff (PAS) reactions, indicating the presence of carbohydrate. It is concluded that CK is a glycoprotein as a carbohydrate, associated with the M subunit of human, rabbit, bovine, and porcine isoenzymes. Also, carbohydrate is an integral part of the B subunit of both rabbit brain and porcine heart CK. Loss of carbohydrate may be important in enzyme catabolism, yielding a pool of circulating modified CK protein that, to date, has remained undetected by traditional methods.
Evaluation of an elsa for the detection of anti‐histoplasma ribosomal and antihistoplasmin antibodies in histoplasmosis
Authors: Raman Chander , Khardori Nancy , Tewari Ram P. , Von Behren Larry A. , Wheat L. Joseph
Published: March 2007

We have developed an indirect sandwich enzyme‐lined immunosorbent assay (ELISA) for the detection of antibodies to Histoplasma ribosomes and histoplasmin; we used this test for demonstration of these antibodies in sera from proven cases of histoplasmosis and other infections. Serum dilutions from five negative controls used in each experiment were normalized against 50 normal sera, and a factor of the mean absorbance was used to establish a positive reaction. Antiribosomal antibodies were detected in 97% of the known histoplasmosis patients with ELISA titers ranging from 1:100 to over 1:12,800. In contrast, antibodies to histoplasmin were detected in only 75% of these sera; titers ranged from 1:100 to 1:12,800. Cross‐reactions with sera from other fungal infections (blastomycosis, coccidioidomycosis, paracoccidioidomycosis, cryptococcosis, candidiasis, and aspergillosis) were seen in 46% of the cases with ribosomes and 37% with histoplasmin. Fifty percent of the sera from tuberculosis patients gave positive reactions with ribosomes and 29% with histoplasmin. These results warrant further studies on the significance of antibodies to ribosomes and histoplasmin in immunity to hlistoplasmosis.
Serum factors in the progression of human immunodeficiency virus type 1 infection to aids
Authors: Tomar Russell H. , John Patricia A. , Hennig Anne K. , Oates Richard P. , Yuille Martin A. R.
Published: March 2007

We studied the prevalence of four serum factors in individuals at different stages of human immunodeficiency virus‐1 (HIV‐1) infection. Soluble interleukin‐2 receptors (slL‐2R) were elevated in all antibody‐positive groups compared with high‐risk, antibody‐negative controls. Paraproteins, usually of the IgG‐kappa isotype, were found in the sera of a significant number of HIV‐1‐infected individuals as were antibodies to lymphocytes (ALAS). Serum factors that inhibit proliferation of peripheral blood mononuclear cells from healthy donors appear late in the course of infection and were associated with increasing clinical severity. Measurement of these factors may prove to be useful in defining the stages of infection and in predicting the appearance or exacerbation of symptoms. They may also play a role in the development of the HIV‐1‐induced immune defects that lead to the expression of clinical acquired immunodeficiency syndrome.
Prostaglandin production in chronic progressive multiple sclerosis
Authors: Aberg Judith A. , Demers Laurence M. , Romano Paula J. , Tenser Richard B.
Published: March 2007

Peripheral blood monocytes have been implicated in the immune reactions that accompany demyelination in patients with multiple sclerosis (MS). We measured prostaglandin E2 (PGE2) and thromboxane B2 (TxB2) release from peripheral monocytes exposed in vitro to complement. Our studies suggest that there is a significantly higher production of PGE2 in monocytes from patients with chronic progressive MS than in those with exacerbation or remitting MS and healthy controls. No significant differences in TxB2release were noted between the three groups.
Simultaneous measurement of neutrophil, lymphocyte, and monocyte glutathione by flow cytometry
Authors: Scott Robert B. , Collin James M. , Matin Sina , White Frances , Swerdlow Paul S.
Published: March 2007

A flow cytometric method for quantitation of glutathione (GSH) was applied to simultaneous analysis of the major leukocyte types in peripheral blood. Cellular thiols (predominately GSH) were stained with monochlorobimane (MCIB), and thiol fluorescence was measured with a flow cytometer. The fluorescence of the thiols closely reflected the GSH content, as measured by a specific glutathione reductase assay. Fluorescence of individual cell types could be measured after delineating those cells by their light‐scatter characteristics, utilizing dual‐angle light scatter for discrimination. By this means, GSH contents of 12.5 ± 2.0nmol/107 neutrophils, 14.5 ± 2.7 nmol/107 monocytes, and 5.0 ± 1.0 nmol/107 lymphocytes were found. The results obtained for neutrophils with the flow cytometer were virtually identical with those obtained with chemical assay in purified samples of neutrophils, indicating the validity of the flow cytometric method.
An elsa procedure to detect anti‐double‐stranded and anti‐single‐stranded dna binding antibodies in connective tissue disorders
Authors: Orr Kenneth B. , Foerster John
Published: March 2007

A simple and sensitive enzyme‐linked immunosorbent assay (ELISA) is described for the measurement of antibodies to native DNA (double‐stranded) and to heat‐denatured DNA (single‐stranded) in sera of patients with connective tissue disorders. DNA bound to polyvinylchloride plates is incubated with serum samples, and antibodies to both single‐ and double‐stranded DNA are detected by means of goat antihuman IgG alkaline phosphatase conjugate. The binding in individual sera is expressed as binding units per milliliter and refers to the absorbance in relation to the absorbance value obtained with a set of standards. The various parameters of the ELISA assay are described, and the results are compared with results of the Crithidia and Farr DNA antibody assays. Two case reports are discussed.
Elevated soluble cd8 in the synovial fluid from patients with rheumatoid arthritis
Authors: Carpenter A. B. , Eisenbeis C. H. , Carrabis S. , Brown M. C. , Ip S. H.
Published: March 2007

Suppressor/cytotoxic T cells express the surface marker CD8, which can be measured in a soluble form in culture supernatants of activated human lymphocytes. Using a sandwich immunoassay, we assessed the levels of soluble CD8 (sCD8) in serum from patients with rheumatoid arthritis (RA; n = 82), patients with degenerative joint disease (DJD; n = 40), and healthy controls. There were no differences in serum sCD8 levels among these groups. In contrast, the levels of soluble CD8 in the synovial fluid (SF) from patients with RA (n = 53) were significantly increased compared with the levels in 23 samples from patients with DJD (821 ± 110 U/ml versus 213 ± 13 U/ml, p < 0.001). Synovial fluid sCD8 levels in the RA group were strikingly elevated, to a maximum value of 5,026 U/ml. In the majority of RA SF specimens (39 of 53), the values were significantly higher in the SF than the serum. Although the RA group had higher values of sCD8, such values were not significantly correlated with measured laboratory or clinical parameters. Current clinical and laboratory methods of evaluating patients may not be adequate in dealing with the complexity and heterogeneity of RA. Soluble CD8 values may be useful in further grouping patients with this disease.
Comparison of prostate acid phosphatase with acid phosphatase isoenzymes from the lung and spleen
Authors: Lin Ming‐Fong , Li Steven S.‐L. , Chu T. Ming , Lee Ching‐Li
Published: March 2007

Acid phosphatase was purified to electrophoretic homogeneity from human normal lung and spleen and was characterized biochemically and immunologically in comparison with prostate acid phosphatase (PAP). The apparent MW of lung acid phosphatase (LAP) and spleen acid phosphatase (SAP) was 110,000 and 100,000, respectively, similar to that of PAP (100,000). All three enzymes exhibited similar electrophoretic mobility, optimal pH, substrate, and inhibitor specificity, except that PAP dephosphorylated profoundly the phosphate group from tyrosine phosphate in phosphoangiotensin (19,700 fmol/mg/min), whereas only marginal activities were detected for LAP and SAP (19 and 73 fmol/mg/min, respectively). Amino acid analysis revealed more similarity between SAP and LAP than PAP and LAP or PAP and SAP. An immunological cross‐reactivity among these three acid phosphatases was detected by polyclonal and monoclonal antibodies raised against purified PAP, although unique epitopes were detected on the PAP molecule. This study provides data explaining why conventional biochemical methods are not specific for PAP measurement and why immunologic methods still detect other acid phosphatases, as observed in clinical laboratory assays. The data also suggest the possibility of using a new substrate or antibody reagent for a more specific assay for PAP.
Measurement of soluble mhc class i molecules in renal graft patients: A noninvasive allograft monitor
Authors: Zavazava N. , Leimenstoll G. , Müller‐Ruchholtz W.
Published: March 2007

We investigated the biochemistry of naturally occurring major histocompatibility complex (MHC) class I molecules in human serum and established a quantitative enzyme‐linked immunoassay (ELISA) to determine soluble human leukocyte antigen (HLA) concentration. Peptides of 46, 40, 37, 35, and 12 kDa were isolated on affinity chromatography columns using two monoclonal antibodies (MoAbs), the anti‐heavy chain W6/32 and the anti‐β‐microglobulin BBM. 1. These peptides were confirmed on Western blots by the HC‐10 MoAb, which binds a monomorphic epitope on denatured heavy chains. In detergent‐binding experiments, only the 46‐kDa peptide could be isolated from solubilized cell membranes. No quantitative differences between serum and plasma HLAs of the same individual were measured. Soluble HLA expression in 12 renal graft recipients was measured over 1–3 mo posttransplantation. A highly significant increase of 50%–100% was noted during rejection episodes. Clinical signs of rejection were accompanied by poor renal function, including elevated creatinine values. After the crises had been managed, class I levels normalized to 0.3–1.5 μg/ml, which is the range in healthy persons. Patients who showed no rejection crises maintained constant levels within the period of study. We anticipate the application of soluble HLA measurement in clinical practice as a noninvasive graft monitor.
Comparison of sex steroid receptor determinations in human breast cancer by enzyme immunoassay and radioligand binding
Authors: Pasic Resad , Djulbegovic Benjamin , Wittliff James L.
Published: March 2007

Most investigators comparing ligand‐binding procedures for quantifying estrogen and progestin receptors in human breast cancer with procedures employing monoclonal antibody‐based methods have utilized an inappropriate variety of reaction conditions, including the elimination of sodium molybdate in the steroid‐binding assays. We studied 197 biopsies of human breast cancer, comparing the results of simultaneous measurements of both estrogen and progestin receptors in identical cytosols by enzyme immunoassay and by radioligand binding using the commercially available kits developed by Abbott Laboratories and by DuPont/NEN Products, respectively. Regression analyses comparing the results from the two procedures indicated a linear relationship, with correlation coefficients ranging from 0.79 to 0.93 for both types of receptors over a wide range of data. Using the widely established cutoff value of 10 fmol/mg of cytosol protein for the ligand‐binding method, and calculating sensitivity and specificity limits according to McNeil et al. (1975), an equivalent cutoff value of 15 fmol/mg of cytosol protein was determined for the enzyme immunoassay of these receptors. Endocrine status of the patient did not appear to alter the cutoff values of either estrogen or progestin receptors when determined by enzyme immunoassay. We recommend that these cutoff values be considered until the results of clinical correlations are completed.
Monoclonal antibody–based immunoassays for serum myoglobin quantification in acute myocardial infarction
Authors: Moscoso Hugo , Kiefer Charles R. , Shyamala M. , Garver Fred A. , Dohlen Von Thomas
Published: March 2007

We have developed a monoclonal antibody–based enzyme immunoassay and a solid‐phase radioimmunoassay for human myoglobin. Both assays are based on competition for the monoclonal antibody between the free myoglobin present in the standards or serum samples and the myoglobin coated to the wells of microtiter plates. Consequently, the absorbance at 630 nm and the radioactivity are inversely related to the concentrations of free myoglobin. The sensitivity of both assays was 10 μg/L with linearity up to 1,000 μg/L. There was no interference with other serum proteins, as judged from analysis of specimens with high concentrations of lactate dehydrogenase, creatine kinase, or hemoglobin. The average serum myoglobin concentration in 30 normal individuals was 67 μg/L. Five patients with cardiac arrhythmias had normal values (average, 63 μg/L) while four patients with myocardial infarction had abnormally high concentrations of myoglobin (300–1,000+ μg/L). In a typical case of myocardial infarction, serum myoglobin rose 21 hr earlier and peaked 12 hr earlier than creatine kinase and its cardiac isoenzyme. These rapid imrnunoassays appear to be useful for the early detection of increased serum myoglobin indicative of myocardial infarction.
Monoclonal free light chains in urine and their significance in clinical diagnostics: Are they really tumor markers?
Authors: Végh Zsuzsa , Ottó Szabolcs , Eckhardt Sándor
Published: March 2007

Bence Jones proteins (monoclonal free light chains of immunoglobulins) are the earliest known biological markers of malignant cell dyscrasia; Bence Jones proteinuria is also present in many types of B cell‐related neoplasms. Sometimes, it may also occur in Hodgkin's disease. In some cases, benign monoclonal gammopathy was found to be associated nontumorous diseases as well. The type of monoclonal light chain, the degree of polymerization, and the isoelectric point of the molecule may affect the course of the disease. Urine samples from 637 patients with true or suspected lymphoproliferative diseases were investigated over a 2‐yr period by different immunochemical methods. Bence Jones proteinuria was identified in 71 cases by isoelectric focusing combined with immunofixation, while the pathological protein was detected only in 63 cases by conventional methods. Bence Jones proteins can be detected by this new method at a level below the sensitivity of conventional procedures. Bence Jones proteins in the urine may signal a malignant tumor or malignant transformation of an earlier disease. The early detection of monoclonal immunoglobulin light chains in the urine may be important in clinical diagnosis, therapy, and follow‐up.
Hepatotoxic effects of repeated administration of oriental hornet (vespa orientalis) venom
Authors: Neuman M. G. , Waron M. , Scapa E. , Eshchar J. , Ishay J. S.
Published: March 2007

Venom sac extract (VSE) of the Oriental hornet (Vespa orientalis) has been shown to exert a toxic effect on hepatic cells. The present work set out to examine its effect on liver cells using serum levels of a lysosomal hydrolase as a marker of injury in rats. Serum activities of beta‐N‐acetyl hexosaminidase (BNAH; EC 3.2.1.30) were found to be positively correlated with the number of treatments received by the rats. The maximal elevation occurred from the third to the sixth day and comprised a 2‐3‐fold increase over the control mean level. This enhancement effect was reversible. The rat envenomend for 2 wk showed 3.5 times as much BNAH activity as in control rats. Biochemical analysis revealed increased specific activity of the lysosomal enzyme in rats treated repeatedly with VSE.
Causitive toxin(s) in the death of two atlantic dolphins
Authors: Hokama Y. , Asahina A. Y. , Hong T. W. P. , Katsura K. , Shang E. , Miyahara J. T. , Sweeney J. C. , Stone R.
Published: March 2007

Following the death of two Atlantic dolphins in a lagoon in March of 1989, the Hawaiian fishes in the lagoon were examined as a potential source of toxin@). This study reports the findings of the causitive toxin(s) involved, utilizing the stick enzyme immuno‐assay (S‐EIA) and the mouse and guinea pig atrium assays. The S‐EIA proved effective in screening the toxic fishes (mullet, wrasse, manini, and aholehole). Following extraction, the major toxin was found in the viscera of these fishes, as confirmed in the mouse assay. The most toxic level was shown in the viscera of the mullet (13.2 mouse units/mg of extract). The viscera of the wrasse, aholehole, and manini also showed high levels of the toxic substance. The guinea pig atrium assay showed the presence of a potent Na + channel inhibitor, characteristic of tetrodotoxin and saxitoxin. The toxin was also demonstrated in low levels in the dolphin liver and gut content and in the sand and algae extracts from the lagoon. This is the first report of this type of toxin in Hawaii.
Detection of hepatitis b virus dna using the polymerase chain reaction technique
Authors: Kaneko Shuichi , Kobayashi Kenichi , Miller Roger H.
Published: March 2007

The polymerase chain reaction (PCR) technique has been utilized for the detection of hepatitis B virus (HBV) DNA, and several factors related to the selection of primer pairs for the PCR amplification have been demonstrated. The sensitivity of the PCR assay was compared with that of slot‐blot hybridization for detecting HBV‐DNA. Analysis by the PCR technique with Southern blot hybridization provided a > 104‐fold increase in sensitivity over the slot‐blot hybridization analysis. Also, a rapid and sensitive PCR method for the detection of serum HBV‐DNA was developed: HBV‐DNA is released from virions by incubating serum with NaOH followed by neutralization with HCI. HBV‐DNA sequences are then detected by agarose gel electrophoresis and ethidium bromide staining after PCR amplification with successive sets of primer pairs. In testing serial samples from chimpanzees experimentally infected with HBV, HBV‐DNA was detected 2–3 wk before the appearance of hepatitis surface antigen (HBsAg) and continued to be detectable for a short period after the production of antibody to HBsAg. Results from testing of human serum demonstrated that the majority of patients with HBsAg in serum had HBV‐DNA as well and that some patients had HBV‐DNA in serum in the absence of HBsAg.
Evaluation of a bench‐top nephelometric immunoassay analyzer
Authors: Van Rijn J. L. M. L. , Van Landeghem A. A. J. , Goldschmidt H. M. J.
Published: March 2007

We evaluated the Technicon DPA‐1 immunoassay analyzer on its analytical characteristics. Therefore we studied three assays: albumin in cerebrospinal fluid and IgG and transferrin in serum. When tested with the Cusum test for linearity albumin, IgG, and transferrin measurements showed no deviation from linearity. Closer examination revealed an abrupt difference of recovery (from 99 to 87%) in the albumin assay when the automatic dilution changed over from the primary analytical to the high analytical concentration range. One calibration was found sufficient for at least 14 days of measurement. Imprecision was well below the critical limits for reproducibility. We found reasonable agreement between the results from the DPA‐1 and the results from comparison methods. However, the correlation plot of IgG showed lack of fit at a distinct segment of the regression line. This appeared to be caused by the poor recovery of the DPA‐1 at the lower limit of the high analytical concentration range. The assays of IgG and transferrin were found insensitive for interference by hemoglobin, triglycerides, urea, and bilirubin. The albumin assay was found sensitive for bilirubin and triglycerides. No reagent‐ and sample‐to‐sample carry‐over could be detected in the assays evaluated.
Monitoring of cyclosporine concentrations by using dry blood‐spot samples
Authors: Mee A. V. , Wong P. Y. , Sun C. , Oei L. , Elliott S. , Naik N. , Joaquin B. , Uchimaru D.
Published: March 2007

We modified the Incstar Cyclo Trac SP kit to enable its use with dry blood‐spots on filter paper. The recovery ranged from 92 to 106%. Dilution studies have shown excellent linearity and parallelism throughout the range of the assay. Precision is demonstrated by within‐assay CV's of 6.6 and 4.3% at 96 and 342 μg/L respectively and between‐assay CV's of 9.1 and 7.0% at 138 and 506 μg/L respectively. A comparison study (n = 209) with whole blood assay gave a correlation coefficient of 0.97, a slope of 1.04, and an intercept of 13.2 Whole blood and dry blood‐spot cyclosporine assays on heart, kidney, liver, and lung transplants were also compared.
Automated immunoturbidimetric analysis of six plasma apolipoproteins: Correlation with radial immunodiffusion assays
Authors: Ikeda Teruaki , Shibuya Yohko , Senba Umeko , Sugiuchi Hiroyuki , Araki Shukuro , Uji Yoshinori , Okabe Hiroaki
Published: March 2007

We measured six apolipoproteins (apo AI, AII, B, CII, CIII, and E) by turbidimetric method using an automatic discrete biochemical analyzer and commercially available antisera. The turbidimetric method was compared with the single radial immunodiffusion method. Linearity for serum apolipoprotein assay by the automated turbidimetric method was better than by the single immunodiffusion method. The linearity by the turbidimetric method was 2.5 G/L for AI, 1.0 G/L for AII, 4.5 G/L for B, 0.12 G/L for CII, 0.3 G/L for CIII, and 0.12 G/L for E. The presence of high concentrations of bilirubin (up to 0.15 G/L) and hemoglobin (up to 50 G/L) interfered with apolipoprotein measurement. Comparison of the immunoturbidimetric and the single radial immunodiffusion (SRID) methods showed excellent coefficients of correlation, r = 0.963, 0.896, 0.846, 0.936, 0.972, and 0.937 for apo AI, AII, B, CII, CIII, and E, respectively.

Reference ranges for the six apolipoproteins were determined by using sera from 450 healthy subjects and were 1.4 ± 0.3 G/L for AI and 0.3 ± 0.01 for E. The observed levels of AII (P <0.001), B (P <0.01), and CIII (P <0.01) were significantly higher in males. The serum levels of apo B, CII, and E showed a gradual increase with age which was more prominent in females than in males. The levels of apo AI, AII decreased significantly over an 11 day period in 22 patients with myocardial infarction.
Isolation of cross‐reacting compounds to incstar cyclo‐trac sp ria in blood samples obtained from cardiac allograft patients on cyclosporine therapy
Authors: Loo Jack C. K. , Gallicano Keith D. , McGilveray Iain J. , Beaudoin Nicole , Jindal Shiv L.
Published: March 2007

To examine the specificity of the Incstar Cyclo‐Trac SP RIA kit, individual blood samples from 28 cardiac allograft patients on cyclosporine A (CsA) therapy were extracted and chromatographed by HPLC. Initially, eluates from a pool of the above samples were collected at regular intervals and measured by RIA to locate possible cross‐reacting compounds. Unknown cross‐reacting materials were detected in a fraction (UNK) that was collected before elution of CsA. For each patient's sample, fraction UNK and the fraction containing CsA were then collected and measured by RIA. In 9 of 28 samples, cross‐reactivity was detected in fraction UNK; range 11 to 36%, mean 22 ± 7.5%. Cross‐reactivity was not apparent in fraction UNK of CsA‐free blood samples from normal volunteers.
Demonstration of human papillomavirus dna by nucleic acid in situ hybridization in paired histologically abnormal cervical biopsies obtained at the same patient visit
Authors: Amortegui Antonio J. , Meyer Michael P. , Kunschner Lara , Saker Adriana
Published: March 2007

Fifty‐four pairs of cervical biopsies ranging from minimal dysplasia to severe dysplasia were studied for the presence of human papillomavirus DNA by in situ hybridization. Two assays were performed on each biopsy. A 16 hour hybridization was used in one assay, while a 40 hour hybridization was utilized in the second assay. Increasing the hybridization time to 40 hours did not significantly increase the detection rate of HPV compared to the rate found using the 16 hour hybridization. Also, no difference in the detection rate of HPV was found by using one biopsy of the pair over the other biopsy of the pair. However, the performance of a single in situ assay on only one biopsy from each patient significantly underestimated the true prevalence of HPV. A single assay only detected 21/33 (64%) patients with HPV. Implications of multiple testing of all histologically abnormal biopsies is discussed in relation to prospective follow‐up studies determining the usefulness of HPV typing in patient management.
Neopterin and β2‐microglobulin and the assessment of intra‐blood‐brain‐barrier synthesis of hiv‐specific and total igg
Authors: Peter James B. , McKeown Karen L. , Barka Noori E. , Tourtellotte Wallace W. , Singer Elyse J. , Syndulko Karl
Published: March 2007

In HIV‐seropositive patients, we evaluated the clinical utility of measuring combinations of serum and CSF levels of neopterin and β2‐microglobulin (β2‐M) (by RIA), as well as the intra‐blood‐brain‐barrier (IBBB) IgG Synthesis rate, IgG index, and HIV antibody index (by rate nephelometry, EIA, and formulae) for the assessment of HIV infection of the CNS. We studied paired sera and CSF from 31 HIV‐seropositive patients: asymptomatic (16), ARC (12), and AIDS (3). A normal serum neopterin level predicts normal levels of serum β2‐M, CSF neopterin, or CSF β2‐M in 90%, 100%, and 100%, respectively, of our patients. An elevated serum neopterin level predicts an elevated level of serum β2‐M or CSF neopterin in 81% and 62%, respectively, of cases. The HIV antibody index and IBBB IgG synthesis rate or IgG index must be determined separately because they do not predict each other and are not predicted by levels of neopterin or β2‐M.
Evaluation of a new western blot strip for human immunodeficiency virus
Authors: Hall Mary F. , Smalley David L.
Published: March 2007

With the isolation of the human immunodeficiency virus (HIV), numerous assays have been developed to detect antibodies to the retrovirus. The manufacturers' recommend that all sera exhibiting a positive enzyme immunoassay be confirmed by the more specific Western blot test (WBT). Recently, a new WBT strip was released for detection of antibodies to HIV. Four hundred patients' sera and 22 CDC proficiency samples were each tested with these strips and with a WBT strip prepared within the laboratory using the CDC techniques. The strips were then evaluated for banding pattern with the reactions recorded on a scale of 0 to 4 + for each band consistent with HIV response. Criteria for positive results were based on the ASTPHLD, CDC criteria. Primary emphasis was on antibody banding to gp41 (envelope protein) and p24 (core protein). The new strips showed appropriate changes in level of antibody, especially for declining antibody to p24. When the banding pattern for antibody to gp41 was compared to the standard strips, the new strips showed a sensitivity of 99.6% and specificity of 99.4%. The presence or absence of antibody to p24 on the new strips versus the standard strips showed a sensitivity of 93.7% and specificity of 99.5%.
Association of serum lipoprotein(a) concentration with other cardiovascular risk factors in a chinese population
Authors: Woo J. , Lam C. W. K.
Published: March 2007

The association between serum Lp(a) concentration, a risk factor for atherothrombotic disease, and other cardiovascular risk factors such as smoking, alcohol intake, hypertension, obesity, and indices of glycemic control is examined in a cohort of 313 elderly Chinese subjects (M = 160, mean age ± SD = 68 ± 11 year; F = 153, mean age ± SD = 73 ± 11). Although associations existed for the above risk factors and other serum lipid levels, there was no association with serum Lp(a) concentration in the overall population. Men with Lp(a) ≥ 30 mg/dl had a higher mean age and blood pressure. A higher percentage of subjects with Lp(a) ≥ 30 mg/dl had a family history of stroke, although the total number of those with a family history was too small to reach statistical significance. The lack of association with known modifiable cardiovascular risk factors is compatible with the conclusion that Lp(a) concentration is primarily under genetic control and therefore unlikely to be a modifiable risk factor.
A filter affinity transfer method for the analysis of rheumatoid factors
Authors: Aucouturier Pierre , Maillochon Jean‐Paul , Joseph‐Théodore Guy , Duarte Francoise , Preud'Homme Jean‐Louis
Published: March 2007

Transfer of serum proteins separated by thin layer agarose electrophoresis onto nitrocellulose sheets precoated with purified human polyclonal lgG followed by revelation with enzyme‐coupled anti‐μ or anti‐α antisera resulted in the specific detection of rheumatoid factors (RF) belonging to the IgM or IgA classes. Mono‐or polyclonality of such RF can be evaluated from the patterns of the blots (sharp bands). In addition, their light chain type can be determined using affinity filters coated with a γ heavy chain disease protein or with IgG Fc fragments. This simple and rapid procedure allows an easy characterization of monoclonal RF, even if they are present in minute amount amongst polyclonal RF as in certain sera from rheumatoid arthritis patients.
Clinical evaluation of five therapeutic drugs using dry film multilayer technology on the opus™ immunoassay system
Authors: Jandreski Mark A. , Shah Jyoti C. , Garbincius Jean , Bermes Edward W.
Published: March 2007

Five therapeutic drug assays, carbamazepine, phenobarbital, phenytoin, theophylline, and valproic acid, were evaluated using an automated random access system for performing thin dry film multilayer competitive immunoassays, the OPUS analyzer. All reagents for the therapeutic drug assays are contained in a coated multilayer film chip encased within a plastic bar‐coded test module and require no external or supplementary reagents. A serum or plasma sample is applied to the test module by the instrument and the fluorescence intensity from the module is measured after 6 minutes. We found the OPUS assays acceptable for clinical use. Within‐run coefficient of variations were 2.3–6.7%, between‐run, 2.9–7.6%. These methods correlated well with the Abbott TDx, having correlation coefficients of 0.92–0.97. Because of the instrument design and the stability of the reagents, weekly calibration is not needed and samples can be run immediately upon receipt in a random access fashion or can be batched together.
Detection of hepatitis C virus (HCV) RNA in paraffin embedded tissue sections of human liver of non‐A, non‐B hepatitis patients by in situ hybridization
Authors: Yamada Shuhei , Koji Takehiko , Nozawa Masayuki , Kiyosawa Kendo , Nakane Paul K.
Published: March 2007

Localization of hepatitis C virus (HCV) RNA was investigated by non‐radioactive in situ hybridization in human liver specimens of chronic non‐A, non‐B (NANB) hepatitis patients who were seropositive for antibodies to HCV (anti‐HCV). For in situ hybridization, T‐T dimerized synthetic oligodeoxynucleotide probes were used and DNAs hybridized in situ were detected immunohistochemically using specific antibodies against T‐T dimer. The data demonstrates that HCV‐RNA was localized in the cytoplasm of hepatocytes in human liver biopsies obtained from the patients with chronic NANB hepatitis seropositive for anti‐HCV.
Reduced levels of free testosterone in four Catania‐type alloalbuminemic males
Authors: Banfi Giuseppe , Daverio Rita , Bonini Pierangelo
Published: March 2007

We Studied total testosterone, sex hormone binding globulin, and free testosterone in four males presenting an electrophoretically slowmoving genetic variant of albumin, the alloalbumin Catania. Free testosterone levels were lower in these cases, found in a year of observation, than those expected for the ages. This finding, which is not related to any disease and constantly not recognized in other males with various genetic variants, should induce consideration of a probable difference of the genetic variant in hormone binding. © 1992 Wiley‐Liss, Inc.
Enzyme immunoassay and immunochemical characterization of pancreatic stone protein in human serum
Authors: Katsuzaki Tomoyuki , Tatemichi Noriyuki , Takeichi Chikako , Hayakawa Shinobu , Hayakawa Tetsuo , Shibata Tokimune , Nakae Yasuyuki , Naruse Satoru
Published: March 2007

Monoclonal antibodies were raised against human pancreatic stone protein (PSP) and used for one‐step enzyme immunoassay (EIA). PSP‐S2‐5 was employed as the standard in the assay. The assay's measurable range was 25‐1,500 ng/ml and within run coefficient of variation was 3.7‐6.4%. Analytical recovery of the assay was 101.5 ± 5.65% (X ± SD). The results of experiments in which serum was fractionated by Mono S (cation exchange chromatography) suggested that most of immunoreactive material in human serum is PSP‐S2‐5. The EIA offers simple, rapid, and specific analysis of serum PSP level for clinical diagnosis. © 1992 Wiley‐Liss, Inc.
Sensitive enzyme immunoassay (immune complex transfer enzyme immunoassay) for (antihuman T‐cell leukemia virus type I) immunoglobulin G in serum using a synthetic peptide, Ala‐Cys‐Env gp46(237–262), as antigen
Authors: Kohno Takeyuki , Sakoda Iwane , Ishikawa Eiji
Published: March 2007

A sensitive enzyme immunoassay (immune complex transfer enzyme immunoassay) for (antihuman T‐cell leukemia virus type l) IgG (anti‐HTLV‐l IgG) in serum using a synthetic peptide, Ala‐Cys‐env gp46(237‐262), of HTLV‐I is described. Anti‐HTLV‐I IgG in test serum, which had been incubated with excess of inactive β‐D‐galactosidase to eliminate interference by anti‐β‐D‐galactosidase antibodies, was reacted simultaneously with 2,4‐dinitrophenyl‐bovine serum albumin‐Ala‐Cys‐env gp46(237‐262) conjugate and Ala‐Cys‐env gp46(237‐262)‐β‐D‐galactosidase conjugate. The complex formed consisting of the three components was trapped onto polystyrene balls coated with affinity‐purified (anti‐2,4‐dinitrophenyl group) IgG. After washing to eliminate nonspecific IgG in the test serum and excess of the β‐D‐galactosidase conjugate, the complex was eluted from the polystyrene balls with ϵN‐2,4‐dinitrophenyl‐L‐lysine and transferred to polystyrene balls coated with affinity‐purified (anti‐human IgG γ‐chain) IgG. β‐D‐galactosidase activity bound to the (antihuman IgG γ‐chain) IgG‐coated polystyrene balls was assayed by fluorometry. This assay was more sensitive than other methods using HTLV‐I antigen, and most negative and positive sera were discriminated. However, some results appeared to be false positive or false negative, and the peptide, Ala‐Cys‐env gp46(237‐262), was suggested to be useful, in combination with other peptides, for improving the reliability of serodiagnosis by separately demonstrating antibodies against as many different epitopes of HTLV‐l as possible. © 1992 Wiley‐Liss, Inc.