bluuebunny/crossref_metadata_2025 huggingface dataset:
Enzyme immunoassay for c‐reactive protein analysis
Authors: Unten Sharon K. , Hokama Y.
Published: March 2007

A solid‐phase enzyme immunoassay method using goat anti‐CRP sensitized to polyvinyl 96 well plates and a second rabbit anti‐CRP highly specific for gamma mobility CRP is described in this study. Examination of serum samples of normal healthy individuals from cord blood, children (2–12 yrs) and adults (16–83 yrs) by this EIA method showed a progressive increase in CRP levels with age. The mean and standard error of the mean in ng/ml CRP was as follows with respect to age distribution: cord blood, 310 ± 54; children (2–12 yrs), 664 ± 87; adults (16–35 yrs), 1,145 ± 116; adults (36–50 yrs), 3,775 ± 605; adults (51–83 yrs), 7,718 ± 1,482. Individuals in the age range of 22–34 yrs with one or more abnormality in the chemistry profile analysis showed CRP levels with a higher x̄ ± S.E.M. (3,615 ± 925). This is in contrast to the normal individuals of the same age range (1,145 ± 116). This EIA method is sensitive, reproducible, and specific. It should be a useful quantitative method for monitoring low level CRP responders such as newborn and cord blood serum.
Solid‐Phase enzyme immunoassay for terminal deoxynucleotidyl transferase (abbott tdt‐eia) in extracts of whole blood and mononuclear cells isolated from bone marrow and blood
Authors: Fairbanks Tod R. , King William J. , Coleman Mary Sue , Finley Paul R. , Fritsche Herbert A. , Hutter John J. , O'Brien John F. , Manderino George L.
Published: March 2007

The Abbott enzyme immunoassay for detecting terminal transferase (TdT‐EIA) was evaluated at four institutions. Extracts of mononuclear cells isolated from bone marrow and peripheral blood of individuals with leukemic and nonleukemic diseases were assayed. In addition, whole blood specimens, in which TdT was extracted from mononuclear cells directly in blood, were assayed. Intrarun, interrun, and between laboratory coefficients of variation ranged from 6.2%–9.3%, 9.4%–11.1%, and 10.9–13.5%, respectively. Since TdT quantitation in whole blood extracts might be adversely affected by blood components and exogenous substances, studies were carried out to determine whether the TdT‐EIA accurately measured TdT in blood. TdT recovery was virtually 100% in blood extracts provided that an extraction buffer, included in the TdT‐EIA kit, was used. Addition of lipids, human gamma globulin, bilirubin, bacterial DNA polymerase, and chemotherapeutic drugs commonly used to treat acute lymphocytic leukemia had no effect on TdT antigenic activity in whole blood specimens. Linear regression analysis at two institutions comparing TdT quantitation using the TdT‐EIA and DNA polymerase assays yielded correlation coefficients of 0.88 and 0.92. Assay results from 1,019 clinical specimens were used to establish expected values for extracts of whole blood and isolated mononuclear cells from blood and bone marrow. The data demonstrate that the TdT‐EIA is a reproducible assay for quantitating TdT levels in clinical specimens.
Twelve‐Protein immunoassay profile on the cobas fara
Authors: Hudson Gail A. , Poulin Sue E. , Ritchie Robert F.
Published: March 2007

The increasing demand for serum protein quantitation poses several problems for the laboratory, including the lack of availability of high‐volume immunochemistry analyzers, the broad concentration range spanned by clinically important proteins, and the difficulty in assay design caused by the variable nature of antiserum. This paper details a profile for 12 commonly requested serum proteins that span a concentration range of three orders of magnitude. The system, which is based on the COBAS FARA centrifugal analyzer, has a capacity of 700 tests per workday while exhibiting performance characteristics that equal or exceed those of pre‐existing techniques. Several features of the FARA and of the assay design that contribute to the high volume capacity are discussed.
Analysis of immunodeficiency in multiple myeloma: Observations and hypothesis
Authors: Pilarski Linda M. , Mant Michael J. , Ruether Bernard A.
Published: March 2007

Patients with multiple myeloma suffer from often profound immunodeficiency that seriously compromises both longevity and quality of life. The cellular basis for the immunodeficiency is unknown and the mechanism(s) giving rise to and possibly maintaining it is also unknown. In this review, evidence defining cellular defects in the nonmalignant B and T peripheral blood lymphocyte pool is summarized and discussed in terms of possible mechanisms that could give rise to the abberrant immune phenotype characteristic of many patients. The abnormalities we have defined include a deficiency of mature B lymphocytes, presence of often large numbers of pre‐B cells in the blood, arrested differentiation of B cells into immunoglobulin (lg)‐secreting cells, and abnormal progenitorlike T cells in blood.

The specificity repertoire of the remaining B lymphocyte population is heavily skewed toward recognition of idiotypic and shared epitopes on the variable region of Ig, perhaps reflecting immune reaction with the monoclonal myeloma Ig. This may begin as a response to tumor antigen that gradually progresses to autoimmunity. A second subset of B cells present at abnormally high frequency in myeloma patients is that of antigen‐experienced memory B cells, which probably encountered antigen prior to the events leading to frank myeloma. In this instance, the high frequency reflects a decrease in aggregate numbers of B cells, although it seems likely that expansion of memory clones also occurs. This is exemplified by the set of memory B cells specific for tetanus toxoid, which are present at normal number but highly enriched frequency in myeloma PBL. Both the antitumor B cells (anti‐idiotype, and anti‐Ig) and the memory cells specific for environmental pathogens appear to be arrested in their differentiation to Ig‐secretors as evidenced by the lack of serum antibody in patients.

The above observations are interpreted in terms of the hypothesis that immunoregulatory events triggered by the monoclonal myeloma Ig establish and maintain the immunodeficiency. We speculate that autoimmune suppressor T cells specific for conserved determinants of Ig prevent pre‐B cell differentiation to slg + B cells, and B cell terminal differentiation to Ig‐secretors. If correct, this analysis could lead to therapeutic approaches for the reversal of the immunodeficiency and perhaps some degree of control over the malignant growth.
Immunoperoxidase staining for calcitonin and calmodulin in the cystic fibrosis lung: A possible new therapeutic strategy in cystic fibrosis
Authors: Wolf Paul , Kilbourn J. P.
Published: March 2007

The presence of calcitonin and calmodulin has been demonstrated by immunoperoxidase staining in formalin‐fixed, paraffinembedded sections of lung from autopsy tissues of patients who died as a result of cystic fibrosis. Calcitonin has been stained and quantitated in solitary endocrine cells, which are increased in number and staining intensity in the cystic fibrosis lung compared to chronic obstructive pulmonary disease (COPD) and normal lungs. Cal modulin was increased in the bronchiolar epithelium and alveolar pneumocytes in cystic fibrosis lung as compared to COPD and normal lungs. Because of our findings, we are proposing a possible new therapeutic regimen for treatment of the pulmonary infection utilizing diuretics and calcium channel blocking agents.
Antibody‐Enhanced enzyme‐amplified determination of methotrexate
Authors: Nashed N. E. , Rechnitz G. A.
Published: March 2007

The sensitivity of potentiometric methotrexate assays can be enhanced by the presence of antimethotrexate antibody, which acts to potentiate the inhibitory effect of methotrexate on enzyme activity. This property is demonstrated in an enzyme‐modified determination of methotrexate with a PCO2 membrane electrode in a cyclic scheme using the enzymes dihydrofolate reductase and 6‐phosphogluconic dehydrogenase.
Homogeneous dry reagent immunoassay strips for the determination of therapeutic drugs in human serum or plasma
Authors: Thompson Stephan G. , Boguslaski Robert C.
Published: March 2007

Specific dry reagent immunoassay strip tests for therapeutic drugs have been developed with the apoenzyme reactivation immunoassay system (ARIS). This simple and convenient method for measuring such analytes in human serum or plasma is described with assays for theophylline, phenytoin, and phenobarbital.
Particle agglutination assay for sm antibodies using purified sm antigen
Authors: Suzuki Hiromasa , Nakamura Robert M. , Tsay Gregory J. , Tan Eng M.
Published: March 2007

Sm antibodies are specific serum markers for the diagnosis of systemic lupus erythematosus (SLE) and are identified by immunodiffusion (ID), counterimmunoelectrophoresis (CIE), and passive hemagglutination (PHA) in the clinical laboratories. These current methods have certain disadvantages of sensitivity, specificity, and complexity.

In this report, Sm antigen free from other nuclear antigens was purified. Synthetic particles were coated with Sm antigen and used to test for Sm antibodies in patient's sera. The particle agglutination test was specific for only Sm antibodies among different antinuclear antisera from patients with systemic rheumatic diseases. The particle agglutination test had the same sensitivity and specificity as the enzyme‐linked immunoassay (ELISA) test for Sm and RNP antibodies. The correlation coefficient between the ELISA quantitation and the titers obtained by particle agglutination was 0.82. The particle agglutination test can be used for rapid screening and titer determination of anti‐Sm antibodies.
Integrated probe for sweat analysis
Authors: Bezegh Klara , Bezegh Andras , Black Philip G. , Janata Jiri
Published: March 2007

Solid‐state integrated probe based on ion‐sensitive, field‐effect transistors has been used for determination of sodium chloride in sweat for diagnosis of cystic fibrosis. This probe can yield useful clinical data faster and with higher reliability than the conventional titration test. A battery‐powered instrument that converts the differential current into milliequivalents of sodium chloride has been constructed. It is possible to perform the diagnosis without inducing sweating with pilocarpine. It is suggested that similar probes with multisensing capability can be used for analysis of other surface body fluids.
Novel enzyme immunoassay of anti‐insulin igG in human serum
Authors: Kohno Takeyuki , Ishikawa Eiji , Sugiyama Satoru , Nakamura Syuji
Published: March 2007

A novel enzyme immunoassay of anti‐insulin IgG in human serum is described. A serum sample containing anti‐insulin IgG was treated with dextran‐charcoal at pH 6.0 to remove endogenous insulin and subsequently incubated with dinitrophenyl biotinyl nonspecific rabbit IgG‐insulin conjugate. The reaction mixture was further incubated with a rabbit (antidinitrophenyl bovine serum albumin) IgG‐coated polystyrene ball to trap the complex formed between anti‐insulin IgG and the conjugate. After washing to eliminate nonspecific IgG in the test serum, the polystyrene ball was incubated with dinitrophenyl‐L‐lysine to elute the complex. The eluate was incubated with an avidin‐coated polystyrene ball. Finally, the amount of human anti‐insulin IgG in the complex trapped onto the avidin‐coated polystyrene ball was measured by incubation with rabbit (antihuman IgG (γ‐chain)) Fab'‐peroxidase conjugate. This enzyme immunoassay was 1,000‐fold more sensitive than the conventional enzyme immunoassay, in which an insulin‐bovine serum albumin‐coated polystyrene ball was incubated with a serum sample containing anti‐insulin IgG and subsequently with rabbit (antihuman IgG (γ‐chain)) Fab'‐peroxidase conjugate. The principle of the novel enzyme immunoassay can be used to more sensitively measure antibodies for most kinds of haptens and antigens than the conventional enzyme immunoassay.
Evaluation of estradiol binding in human benign prostatic hyperplasia
Authors: Ganesan S. , Bashirelahi N. , Young J. D.
Published: March 2007

We have systematically evaluated the effect of tris/phosphate buffers, ethylenediamine tetraacetate (EDTA), dithiothreitol (DTT), molybdate, and phenylmethylsulfonyl fluoride (PMSF) on the binding of 17β‐estradiol to human BPH cytosol. Competition binding assays demonstrated that the estrogen receptor was highly specific for 17β‐estradiol. There was statistically no difference in binding observed with tris or phosphate buffers. Two classes of binding (high affinity—type I, low affinity—type II) were observed. DTT (1 mM) showed maximum inhibition of type II binding. There was no significant difference in binding observed in the presence of 1.0 mM EDTA. However, 10 mM EDTA stimulated estradiol binding in both saturation analysis and glycerol gradients. Stimulation of estradiol binding by 10 mM EDTA was observed even in the presence of 1 mM DTT. Molybdate (20 mM), a reagent used to stabilize steroid receptors, significantly inhibited estradiol binding. PMSF inhibited estradiol binding at all concentrations studied and the nature of the inhibition appeared to be uncompetitive. A Bmax (maximum number of binding sites) of 84.4 ± 68.5 fmoles/mg protein with a KD (equilibrium dissociation constant) of 2.5 ± 1.7 mM was observed for type I estrogen receptor binding. Our Bmax values are higher than those reported in other studies even after allowing for possible overestimation. In view of the potential importance of estrogen receptors in human prostate, careful reevaluation of estrogen receptors must be undertaken using proper assay conditions.
ELISA assay for colon cancer
Authors: Lawless B. Desales , Lee Yu‐Chen C. , Crook James E. , Clapp Neal K.
Published: March 2007

The high mortality rate for patients with colon cancer results largely from the inability to diagnose the condition in its early stages. The assay described in this report may be useful in early detection of colon cancer. In the study we use the cotton‐top tamarin, Saguinus oedipus, as an animal model for studying human colon cancer. This New World monkey develops a high incidence of colitis and colon cancer. Antibodies that recognize adenocarcinoma cells and antigens from these cells can be detected in the sera of tamarins with colon cancer. The antibodies are not found in sera of healthy animals. In one procedure, tamarin adenocarcinoma cells are used to coat the wells of a microtiter plate. In an alternate procedure, a glycoprotein adenocarcinoma antigen is used. The antigen is extracted from a cultured tamarin colon cancer line by detergent solubilization followed by affinity chromatography. Test sera are examined by adding biotinylated immunoglobulin serum fractions to the coated wells. The wells are washed and an avidin‐enzyme conjugate is added. Positive sera are identified by the addition of the substrate. The presence of the autoantibodies in the sera may be taken as evidence that the animal has colon cancer.
Current status of fluorescent immunoassays
Authors: Robbins Bruce A. , Nakamura Robert M.
Published: March 2007

In 1941 fluorescent compounds were first introduced as immunochemical labels in assay systems. Several forms of quantitative fluorescent immunoassay, which represent a major alternative to radioimmunoassay and enzyme immunoassay, have been developed and are now widely used in many clinical laboratories. This paper discusses these forms and procedures and their advantages over radioimmunoassay and enzyme immunoassay.
Biochemical nature and biological activity of the vascular permeability factor of bovine platelets
Authors: Ashihara Yoshihiro
Published: March 2007

Bovine vascular permeability factor (BVPF) was purified from the α‐granule acid extract of platelets (BPAE) by ion‐exchange and gel‐filtration chromatography. BVPF is a cationic protein with an isoelectric point of 8.8. As determined by gel filtration using a Superose 12 column, there appear to be three kinds of BVPF with molecular weights of 30,000, 300,000, and > 450,000. After heat treatment of BPAE, however, BVPF was eluted in a fraction with a molecular weight of 30,000. Vascular permeability increased as much as 2.3‐fold with heat treatment of BPAE at 56°C for 30 minutes and was retained even after heat treatment at 80°C. BVPF was also stable to acid or alkaline treatment, but it was sensitive to proteases or thiol reagents, especially pepsin and 2‐mercapto‐ethanol. After injection of BVPF into the dorsal skin of rabbits, enhancement of vascular Permeability was reconfirmed by 3 minutes of observation electron microscopically. One hour later, exudation had increased at the injection sites, and there was associated inflammatory cell infiltration with occasional strands of fibrin.
Potentiometric determination of serum 5′‐nucleotidase
Authors: Ho May Y. K. , Rechnitz Garry A. , Chen Ted K.
Published: March 2007

An ammonia gas sensor was used to determine 5′‐nucleotidase in serum through the ammonia produced from the coupled reactions of 5′‐nucleotidase and adenosine deaminase. Nonspecific hydrolysis of the substrate 5′‐adenosine monophosphate by serum alkaline phosphatase was corrected for by using the inhibitor concanavalin A. The present method is fast and simple, and offers considerable improvements over existing methods. Results obtained using the present method compare well with the spectrophotometric method and are not affected by sample turbidity.
Correlation of serum levels of CA‐549 to disease status in posttreatment serial samples from breast cancer patients
Authors: Bray Kurtis R. , Gaur Pramod K.
Published: March 2007

CA‐549 is a high molecular weight, circulating marker for breast cancer defined by its reactivity to monoclonal antibody BC4E 549. CA‐549 has recently been shown to be elevated in the sera of 50% of advanced breast cancer patients (n = 80), as compared to <2.0% of benign breast disease patients (n = 79) or healthy individuals (n = 257).

The purpose of this study was to evaluate CA‐549 as a tool for monitoring the status of breast cancer disease. Serial serum specimens (n = 381) from a total of 126 metastasis breast cancer patients were prospectively collected over 12 months and tested for CA‐549 levels by immunoradiometric assay. The average percent increase in CA‐549 levels in the 18 patients with disease progression was 115 ± 103% (range = −27% to 440%), with 16 of 18 showing an increase of >25%. CA‐549 levels decreased by a mean of −40.5 ± 22.5% (range = 17% to −75%) in 12 patients with breast cancer regression, with 8 of 12 patients showing a decrease in serum CA‐549 of >25%. For 18 patients diagnosed with clinically stable disease, the mean percent change in CA‐549 was only 20.4 ± 5.7%, range = (−27% to 24%), with 14 of 18 having changed by <25%. For patients with no evidence of disease, 75 of 78 were CA‐549 negative throughout the study. Three of 78 patients showed borderline positive CA‐549 elevation for at least one specimen. None of these 78 patients showed highly elevated CA‐549.

This study confirms the highly specific nature of CA‐549 as a breast cancer marker, and indicates that changes in CA‐549 levels are associated with the clinical course of the disease. Thus this test may be a useful tool to monitor response to treatment and recurrence of breast cancer.
Enzyme inhibitory homogeneous immunoassay for high molecular weight antigen (I)
Authors: Ashihara Yoshihiro , Nishizono Isao , Tanimoto Tetsuji , Tsuchiya Hiromichi , Yamamoto Katsuhiko , Kido Yasuji , Miyagawa Eiji , Kasahara Yasushi
Published: March 2007

A highly sensitive homogeneous enzyme immunoassay has been developed for the determination of antigens with high molecular weight. The method presented is based on steric hindrance of the enzyme against an insoluble substrate by antigen binding to antibody: enzyme, conjugate, and consists of α‐amylase from Bacillus subtilis as label with antibody, human α‐fetoprotein (AFP) or human ferritin as ligands, and insoluble substrate. The assay procedure is as follows: sample is mixed and incubated with the conjugate at 37° C for 20 min. Thereafter, the mixture is incubated at 37° C for 20 min following addition of substrate, and the enzyme reaction is terminated by adding the stopper. Then the mixture is centrifuged at 3,000 rpm for 1 minute, and the absorbance of the supernatant is measured at 620 nm. The measurable range for AFP was 5‐200 ng/ml for ferritin, it was 10‐800 ng/ml. In addition, the method was well correlated with radioimmunoassay; i.e., for AFP r = 0.892 and for ferritin r = 0.951. Furthermore, there was no interference during the assay of endogenous a‐amylase in human serum as a result of the use of the inhibitor specific to mammalian amylase which was purified from bacteria.
C‐reactive protein and platelet activating factor complexes induce production and release of interleukin‐1 by human monocytes
Authors: Hokama Y. , Honda S. A. A. , Hanakahi L. A. , Terada K. Y.
Published: March 2007

Stimulation of interleukin‐1 (IL‐1) in peripheral blood monocytes exposed to C‐reactive protein (CRP) and C‐reactive protein‐platelet‐activating factor (CRP‐PAF) complexes was demonstrated in vitro in this study. Significant synthesis of intracellular IL‐1, 2‐5‐fold increases in the number of adherent monocytes as determined by indirect immunofluorescence using rabbit anti‐IL‐1 and goat antirabbit IgG‐fluorescein isothiocynate, was shown. Both CRP alone (10 μg/ml) and especially CRP‐PAF mixture (10 μg CRP/1 ug PAF/ml) showed maximum stimulation of IL‐1 of 41 and 95 cells/10 field examined, respectively. CRP alone examined in a dose‐response study showed optimum stimulation of IL‐1 production at the 1 μg/ml concentration with 57% of monocytes showing fluorescence. CRP at varying concentrations with a constant amount of PAF (1 μg/ml) showed optimum IL‐1 staining monocytes (63%) with CRP at 10 μg/ml (10 pg CRP/l μg PAF). Similar significant enhanced extracellular IL‐1 production by monocytes exposed to CRP and CRP/PAF mixtures was shown by the mouse thymocyte assay. Supernatants of monocytes exposed to CRP (10 μg/ml) or CRP‐PAF (10 μg/l μg/ml) demonstrated mouse IL‐1 synthesis of 92 and 110 stimulation indices in the thymocyte assay, respectively. It is suggested that CRP and especially CRP‐PAF complexes activate macrophage through their appropriate receptors to production of monokines which modulate the immune system. Thus, CRP appears to function nonspecifically in a behavior reminiscent of specific immunoglobulins.
CA‐549: A new tumor marker for patients with advanced breast cancer
Authors: Demers Laurence M. , Harvey Harold A. , Glenn Jerry D. , Gaur Pramod K.
Published: March 2007

A murine monoclonal antibody, BC4E 549, an IgG1 adapted as tracer antibody in a sandwich immunoradiometric assay, was used to detect a circulating breast cancer‐associated antigen, CA‐549, in patients with advanced breast cancer. The study was designed to examine the specificity and sensitivity of CA‐549 measurements as a specific circulating marker for breast cancer. In a population of 100 normal pre‐and postmenopausal women, serum levels ranged from 1–10 μ/ml. In contrast, 38 of 46 patients (83%) with active, advanced‐stage breast cancer had circulating levels in excess of 10 μ/ml, and levels in several patients exceeded 100 μ/ml. Patients on adjuvant therapy with breast cancer in remission, patients with benign breast disease, and patients with cancers of other sites such as colorectal, ovarian, and lung cancer did not show significant elevations (>10 μ/ml, P > 0.001) of this circulating antigen. Measurement of this breast cancer associated antigen may be of use in the evaluation of treatment for patients with advanced stage breast cancer.
Serum levels of cancer‐associated antigen 195, a circulating marker for colon cancer, and its relationship to carcinoembryonic antigen
Authors: Bray Kurtis R. , Gaur Pramod K.
Published: March 2007

CA‐195 is a circulating cancer‐associated antigen defined by its reactivity with monoclonal antibody CC3C 195. The epitope with which this antibody reacts has previously been shown to be the Lewis A blood group antigen and its sialylated derivative.

By using CC3C 195 as both capture and tracer antibody, serum levels of CA‐195 were measured with an immunoradiometric assay. CA‐195 was found to be elevated in the sera of 36 of 68 advanced colon cancer patients, 12 of 26 advanced liver cancer patients, 9 of 30 advanced breast cancer patients, and 4 of 19 advanced lung cancer patients. This is in contrast to healthy individuals, in which only 11 of 203 had elevated CA‐195, and to benign colon disease patients, which had 10 of 76 elevated. Benign breast disease patients, benign liver disease patients, and benign lung disease patients had 5 of 30, 6 of 25, and 2 of 30 sera with elevated CA‐195 levels, respectively. This elevation of serum CA‐195 was statistically significant in colon cancer patients compared to benign colon patients (P<.0001) or healthy individuals (P<.0001), as well as in liver cancer patients compared to benign liver patients (P < .025). Elevations in other conditions studied were not statistically significant. A separate panel of sera from staged, pretreatment colon cancer patients was also tested, and elevated CA‐195 was found in 6 of 17, 17 of 54, 10 of 18, and 12 of 17 patients which were staged Dukes A, 8, C, and D, respectively. Dukes C and/or D patients were significantly elevated compared to Dukes A and/or 6 (P<.0035), and all stages were significantly elevated compared to sera from healthy individuals (P<.0001). This indicates that the incidence of CA‐195 elevation increases as the disease progresses and suggests that CA‐195 may be a useful marker at all stages. Anticarcinoembryonic antigen (CEA) and anti CA‐195‐coated beads were used to immunodeplete specimens; CA‐195 epitopes were found to be present on the same molecular structure as CEA epitopes in tumors but not in patients' sera. This confirms the nature of CA‐195 as a separate cancer marker in patients' sera and suggests that cleavage of CA‐195 and CEA epitopes from a larger precursor moiety occurs prior to secretion into the serum.
Dot‐immunobinding assay with the globular domain of collagen type IV for antiglomerular basement membrane antibodies
Authors: Weber M. , Manns M. , Meyer zum Büschenfelde K.‐H. , Köhler H.
Published: March 2007

A dot‐immunobinding assay for the detection of antiglomerular basement membrane antibodies has been developed. The globular domain NC1 of basement membrane collagen type IV was used as antigen. The assay proved to be specific, sensitive, and reproducible. Circulating antibodies in each of 12 sera from patients with florid Goodpasture's syndrome could be demonstrated, whereas sera from patients with Goodpasture's syndrome in clinical remission and various control sera showed no reactivity. The advantages of the dot‐blot assay are: the usage of the purified Goodpasature target antigen NCI reduces unspecific binding of IgG; only minimal amounts of antigen are required to give a positive signal; evaluation of the assay is possible without expensive laboratory equipment; precoated nitrocellulose sheets can be stored for several months without loss of activity.
IgA With altered carbohydrate structure as a tumor‐associated marker in serum of cancer patients
Authors: Henslee Jerry G. , Rittenhouse Harry G. , Matias Matthew S. , Michael Hass G. , Schenck Jay R. , Tomita Joseph T.
Published: March 2007

Serum IgA1 contains O‐linked carbohydrate structures, whereas other immuno‐globulins contain only N‐linked structures. An assay was developed which detects IgA1 by measuring the DGalβ(1–3) DGalNAc structures O‐linked to the IgA1 hinge region. Fifty percent of the serum specimens from cancer patients tested were elevated compared to 1% of the total normal serum specimens tested by this assay. The assay combines antibody (anti‐human IgA) and lectin and measures the DGalβ(1–3)DGalNAc structure in the IgA1 molecule accessible to lectin binding. Three lectins were compared–the lectins from Bauhinia purpurea alba, Maclura pomifera, and peanut agglutinin (PNA). PNA in the assay discriminated best between serum specimens from normal subjects and cancer patients and was chosen as the lectin component in the assay. The anti‐IgA‐PNA enzyme immunoassay was evaluated in a retrospective study of cancer patients. A panel of 845 serum specimens was tested and the results analyzed at three cutoff R values. The R value of a specimen is the ratio of the assay absorbance obtained for that test specimen with respect to the assay absorbance obtained for a pool of normal human sera. At a cutoff R value of 2.38, 1% normals and 26% benigns were elevated. The following cancer specimens had elevated R values: 52% lung, 60% colon, 43% head and neck, 35% breast, 64% kidney, 38% bladder, 43% prostate, 8% ovarian, and 67% pancreas. Further testing showed a significant percentage of serum specimens from patients with cirrhosis, pancreatitis, or ulcerative colitis also had elevated R values. No correlation of this assay was observed with CEA levels. The results suggest that the carbohydrate structure of serum IgA1 is altered by malignancies and that measurement of this altered IgA may supplement CEA testing.
Sandwich immunoassay for the detection of lipoteichoic acid
Authors: Tadler Monica B. , Tice George , Wos Susan M.
Published: March 2007

A sandwich immunoassay has been developed for the detection of lipoteichoic acid (LTA), a major cell wall constituent of gram‐positive bacteria, from whole blood and ISOLATOR supernate. Monoclonal antibodies were produced to purified LTA from Streptococcus mutans, BHT and were further characterized for crossreactivity with gram‐positive and negative bacteria and for reactivity to substituted and unsubstituted LTA. Eight monoclonal antibodies were identified that reacted exclusively with gram‐positive bacteria. Those antibodies able to capture H‐LTA were chosen to develop a sandwich immunoassay. The assay has a sensitivity of 0.2 ng LTA/mL in PBS, 0.5 ng/mL in whole blood and 2.0 ng/mL in whole blood that has been processed through the ISOLATOR. Further development of this assay may lead to the rapid detection of LTA from body fluids.
Influence of methisoprinol (lsoprinosine) on hiv‐infected human lymphocytes: In vitro immunological, virological, and ultrastructural studies
Authors: De Simone Claudio , DeMarco Federico , Arancia Giuseppe , Tzantzoglou Sonia , Paradisi Silvia , Sorice Franco
Published: March 2007

Methisoprinol (isoprinosine) is a synthetic compound with reported antiviral and immunomodulating properties. Results of the present study showed that methisoprinol at concentrations ≥ 200 μg/ml reduces the p24 and gp120 viral antigen expression on the surface of human immunodeficiency virus (HIV)‐infected lymphocytes and the reverse transcriptase levels. In addition, cell viability, the number of the CD4 + cells, and the CD4 + /CD8+ cell ratio are higher in methisoprinol‐pretreated cell suspensions than in untreated HIV‐infected cell cultures. A quantitative freeze‐fracture study on the density of the intramembranous particles (IMP) present on both fracture faces of the plasma membrane of lymphocytes has shown that pretreatment with methisoprinol induces a different molecular organization resulting in a nearly three times increase of IMP density.
Human monoclonal antibodies reactive with colon carcinoma: Identification by a novel screening procedure with xenografts
Authors: Kan‐Mitchell June , Formenti Silvia C. , Mitchell Malcolm S.
Published: March 2007

To identify tumor‐associated antigens immunogenic to man, human monoclonal antibodies (MAbs) were generated by fusing regional lymph node lymphocytes (LNLs) of each of three colon adenocarcinoma patients with the M5 mouse myeloma cell line. Heterohybridomas in 19 wells secreted human immunoglobulin (lg) for at least 60 days. To identify immunoreactive MAbs, we devised an immunohistochemical assay with xenografts of six colon tumors. Binding was visualized with a biotin‐conjugated goat antihuman lg antibody followed by an avidin‐biotin‐peroxidase complex. This assay detected binding with a sensitivity of 0.5 ug/ml human lg. Reactivity against tumor‐associated antigens (TAAs) in frozen sections of the colon xenografts was detected in 14 of the 19 supernates. However, only nine (three IgGs, five lgMs and one IgA) were reactive against formalin‐fixed sections, showing that formalin‐fixation destroyed some but not all antigenicity. Four MAbs were cloned and tested against the entire panel of 19 xenografts. One IgM MAb reacted only against colon carcinomas, while the other three (one each of IgA, IgG and IgM) had broader reactivities. Three of the four human MAbs did not cross‐react with melanomas. In summary, human regional lymph nodes contain a high proportion of B lymphocytes sensitized to the autochthonous tumor. Immunostaining of human colon tumor xenografts can readily identify human MAbs reactive with colon carcinomas.
Analysis of peroxidase negative acute leukemias by monoclonal antibodies: III. Acute lymphoblastic leukemia
Authors: Imamura Nobutaka , Kuramoto Atsushi
Published: March 2007

Peripheral‐blood leukemic cells from 45 patients with peroxidase negative acute lymphoblastic leukemia (ALL), which did not express either myeloid or megakaryocyte‐platelet‐related cell surface antigens, were analyzed by using monoclonal antibodies capable of recognizing T‐ or B‐cell‐associated and/or T‐ or 6‐cell‐restricted antigens. Numerous subclasses of ALL, including B‐cell lineage leukemias and T‐cell lineage leukemias, were identified phenotypically and immunophenotypically in an effort to more accurately characterize the heterogeneous ALLs, their states of differentiation, and their relationships to normal B‐ and T‐lymphoid cells. Among the cases studied, only seven (15.6%) were found to have stem cell (undifferentiated) leukemia (la+, CD24+, CD9+, CD34+). It is concluded that the use of monoclonal antibodies for the characterization of heterogeneous ALLs improves the specificity of leukemia classification, which may contribute to the selection of more effective forms of therapy for the types of leukemia identified.
Characteristics and distribution of receptors for [d‐trp6]‐ luteinizing hormone‐releasing hormone, somatostatin, epidermal growth factor, and sex steroids in 500 biopsy samples of human breast cancer
Authors: Fekete Matyas , Wittliff James L. , Schally Andrew V.
Published: March 2007

Binding capacities and apparent dissociation constants of receptors for [D‐Trp6]‐luteinizing hormone‐releasing hormone ([D‐Trp6]‐LH‐RH), somatostatin (SS‐14), epidermal growth factor (EGF), and estrogen and progesterone were determined in 500 breast cancer specimens using multipoint assays. Specific binding sites greater than 10 fmol/mg cytosol protein for estrogen were found in 408 carcinomas (81.6%), and for progesterone in 340 specimens (68%). High affinity EGF receptors were present in membrane preparations from 335 samples (67%). In 260 of 500 samples (52%), two classes of [D‐Trp6]‐LH‐RH membrane receptor sites were also detected, one class showing high affinity and low capacity, and the other class showing low affinity and high capacity; 178 biopsy samples (35.6%) exhibited binding sites for SS‐14. Statistically significant inverse correlations were found between the binding capacities of estrogen and EGF receptors as well as between Bmax of progesterone and EGF receptors. Significant positive correlations were demonstrated between binding capacities of estrogen and progesterone and between Bmax of high affinity and low affinity binding sites of [D‐Trp6]‐LH‐RH receptors. However, no correlation was found between the dissociation constants of different receptor sites in human breast cancer specimens. These results demonstrate that numerous human breast cancers, in addition to receptors for estrogen and progesterone, also show binding sites for EGF, [D‐Trp6]‐LH‐RH and SS‐14. The methods described herein permit a routine quantification of receptor sites for [D‐Trp6]‐LH‐RH, SS‐14, and EGF in membrane preparations of biopsy samples of breast cancer and can be used in conjunction with the determination of estrogen and progesterone receptors in nuclear‐cytosolic extracts. The simultaneous measurements using a microanalytic approach allow the determination of peptide and steroid hormone receptors that might be involved in the response mechanisms of human breast cancer. It should be possible to correlate the levels of these receptors with clinical parameters to better identify endocrine‐responsive neoplasms. This approach might be useful to guide a rational hormonal therapy in women with breast cancer.
IgM to human cytomegalovirus: Comparison of two enzyme immunoassays and igm reactivity to viral polypeptides detected by immunoblotting
Authors: Re M. C. , Landini M. P.
Published: March 2007

IgM detection by enzyme‐linked immunosorbent assay (ELISA) is the most used method of establishing a cytomegalovirus (CMV) infection during pregnancy. In this paper, we discuss the results obtained assaying 1,000 sera from pregnant women by two ELISA kits: a traditional indirect ELISA and a more recent IgM‐capture ELISA. All the sera that gave a positive ELISA value with one or both kits were further tested by immunoblotting (IB) to establish which CMV polypeptides were detected by IgM antibodies. From the results obtained, IgM‐capture ELSA seems less sensitive than indirect ELISA, but correlates better with serological evidence of active CMV infections as judged by typical IB profiles.
Detection of a tumor‐associated glycoprotein antigen in serum and urine of melanoma patients by murine monoclonal antibody (AD1‐40F4) in enzyme immunoassay
Authors: Euhus David M. , Gupta Rishab K. , Morton Donald L.
Published: March 2007

The urine of 68% of melanoma patients contains a high molecular weight glycoprotein which is expressed by melanoma cells and reacts with autologous antibody. Since high levels of this antigen in urine correlate with disease recurrence in surgically treated melanoma patients, it has been termed urinary tumor‐associated antigen (U‐TAA). We report the development of a murine monoclonal IgM antibody (AD1‐40F4), which is specific for U‐TAA. AD1‐40F4 showed the same pattern of reactivity as the allo‐antibodies previously used for the detection of U‐TAA. The antigen recognized by AD1‐40F4 has a high molecular weight (590– 620 kilodaltons [kd]) and is heat stable. The AD1‐40F4‐reactive epitope is a protein. When AD1‐40F4 was applied in an enzyme immunoassay, it allowed for the detection of U‐TAA in the serum of 64% (33/52) of melanoma patients as opposed to only 5% (1/20) of normal controls. Thus, the murine monoclonal antibody AD1‐40F4, which has been specifically developed against an allogeneic antibody defined antigen, U‐TAA, appears to be important for immuno‐prognosis of human melanoma.
Effect of azidothymidine (AZT) on P24 antigen levels in patients with AIDS‐related complex and AIDS
Authors: Reddy Mohan M. , McKinley George , Englard Arthur , Grieco Michael H.
Published: March 2007

Circulating HIV P24 antigen levels at baseline and following AZT therapy were measured in 9 patients with ARC and 11 patients with AIDS. Eight of these patients had no detectable levels of P24. The P24 antigen levels in the remaining 12 patients decreased significantly after AZT treatment. Following discontinuation of AZT treatment in 2 patients, the P24 antigen levels went up. These results clearly demonstrate the efficacy of AZT in reducing P24 antigen levels in circulation.
Effect of a marine toxin on human peripheral blood monocytes
Authors: Hokama Y. , Scheuer P. J. , Yasumoto T.
Published: March 2007

The effect of okadaic acid (OA), a non‐TPA (12‐0‐tetradecanoylphorbol‐13—acetate)‐type tumor promoter, on interleukin 1 (IL‐1) synthesis in human peripheral blood monocytes in vitro was examined using immunofluorescence and the mouse thymocyte assays. Stimulation of IL‐1 was shown with 0.05 μg OA/ml (the peak response), while concentrations of greater than 1.0 μg OA/ml showed significant inhibition of IL‐1 synthesis by monocytes in the immunofluorescence analysis. Okadaic acid added directly to mouse thymocytes showed a peak response in IL‐1 synthesis at 0.01 μg OA/ml, while significant inhibition was shown for concentrations of OA greater than 0.1 μg ONA/ml. Supernatants of monocytes exposed to OA gave maximum stimulation at 0.05 μg OA/ml for both 24 and 48 hr exposures. Significant inhibition of IL‐1 synthesis in monocytes was shown by supernatants obtained from monocytes exposed 24 hr with 1.0 μg OA/ml. Addition of various concentrations of OA to monocyte cultures in the presence of increasing concentrations of monoclonal antibody to OA showed significant reduction of the inhibition of IL‐1 synthesis by OA at the 0.10 μg level, but more significantly at the 1.0 μg OA/ml level. The higher levels of OA associated with inhibition of IL‐1 synthesis in monocytes in this study were comparable to the concentrations used in the tumor promotion studies by others. The reversal of the inhibitory effect of OA by the monoclonal antibody to OA is of interest and should be applicable to further studies on the mechanism of OA tumor promotion.
Jacalin: A new laboratory tool in immunochemistry and cellular immunology
Authors: Aucouturier Pierre , Pineau Nathalie , Brugier Jean‐Christophe , Mihaesco Edith , Duarte Françoise , Skvaril Frantisek , Preud'homme Jean‐Louis
Published: March 2007

In recent years, a new lectin—jacalin—has raised the interest of immunologists because of its original properties with respect to human immunoglobulins and lymphocytes. Its structure and carbohydrate binding specifity are now well documented, and it can be purified easily from jackfruit seeds by ion exchange or affinity chromatography. The binding and precipitating specificities of jacalin with heavy chains of human immunoglobulins allow its use as a diagnostic (IgA subclass typing) and preparative tool (purification of IgA and IgD, removal of IgA from biologic samples and preparations). Other possible applications of jacalin's binding properties also can be envisaged. In addition, the lectin displays a mitogenic activity specific for human CD4 T‐lymphocytes; consequently, the proliferative response induced by jacalin appears to represent a new and interesting assay for a functional study of CD4 cells, with obvious applications in primary and acquired, especially AIDS, immune deficiency states.
Comparison of igg subclasses and complement binding activity of autoantibodies from patients with bullous pemphigoid and pemphigus
Authors: Brooks Walter S. , Lee Yue‐Yun , Abell Edward , Deng Jau‐Shyong
Published: March 2007

Bullous pemphigoid and pemphigus are autoimmune disorders of skin of unknown etiology and are characterized by the presence of immunoreactants in the skin and circulating autoantibodies to skin components. The distribution of IgG subclass antibodies to intercellular substance (ICS) of pemphigus and basement membrane zone substance (BMZ) of bullous pemphigoid was analyzed by using monoclonal antibodies to human IgG subclasses. IgG, type anti‐BMZ antibody was found in the majority of patients with bullous pemphigoid (88% in skin and 96% in serum). One third to one half of bullous pemphigoid patients had lgG, and IgG, anti‐BMZ antibodies. The majority of bullous pemphigoid skin (92%) had complement in skin, however only one third of their sera had complement binding activity in vitro. lgG, anti‐ICS antibody was the predominant one in patients with pemphigus (86% in skin and 80% in circulation). IgG4 anti‐ICS antibody was seen in two thirds of specimens from pemphigus patients. IgG, subclass antibody was more frequently seen in pemphigus than in bullous pemphigoid patients. Two‐thirds of pemphigus sera were capable of activating complement in vitro. The complement binding activity was directly associated with IgG1 and/or IgG3 subclass antibodies. The possible mechanisms for the restricted IgG4 subclass antibodies in bullous pemphigoid and pemphigus are discussed.
Effects of substance p on the spontaneous binding of salmonella minnesota r345 (rb) to human peripheral blood lymphocytes
Authors: De Simone Claudio , Misefari Aldo , Antonaci Salvatore , Maffione Angela B. , Jirillo Emilio , Covelli Vito
Published: March 2007

The effects of substance P (SP) on Salmonella minnesota R345 (Rb) binding to human peripheral blood lymphocytes (PBL) were evaluated. Two parameters of bacterial cytoadherence were considered, namely the binding lymphocytes (BL) and the number of bound‐bacteria/lymphocyte (BB). The results showed that SP inhibits both BL and BB in a significant manner. Furthermore, distribution of Salmonella binding to CD4+ and CD8 + lymphocytes was studied following SP pre‐treatment of lymphoid cells. This neuropeptide is able to hamper the bacterial cytoadherence to both T‐cell subpopulations and, in particular, the inhibitory effect on the T‐suppressor/cytotoxic subset was more pronounced.

These findings are discussed in terms of SP intervention in the mechanism of host protection against invading microorganisms.
Serum levels of interleukin‐2 receptor and disease activity in patients with iga nephropathy
Authors: Tomino Yasuhiko , Ozaki Teruyo , Koide Hikaru , Takahashi Masayoshi , Ito Kiichi
Published: March 2007

The levels of serum interleukin‐2 receptor (IL‐2R) were studied by enzyme‐linked immunosorbent assay (ELISA) in patients with IgA nephropathy. The aim of the present study was to determine if levels of serum IL‐2R might correlate with disease activity in patients with IgA nephropathy. Twenty‐eight patients with IgA nephropathy were examined.

This study showed a significant correlation between the levels of serum IL‐2R and disease activities, i.e., levels of urinary protein, blood urea nitrogen (BUN) and uric acid, in patients with IgA nephropathy. The levels of serum IL‐2R in patients with moderate or advanced stage of IgA nephropathy were significantly higher than those in patients with the minimal or slight stage of this disease or in healthy adults. It was suggested that the measurement of serum IL‐2R is useful in evaluating the degree of disease activity and/or prognosis in patients with IgA nephropathy.
Human psychoneuroimmunology today
Authors: Biondi Massimo , Kotzalidis Giorgio D.
Published: March 2007

Studies in human psychoneuroimmunology began around 1919, but a systematic approach wasn't used until the work of Solomon in the 1960s. Recently, the new specialty has achieved relative independence due to considerable data acquisition.

Stress research has revealed relationships between neuroendocrine and immune changes. In parallel, increasing evidence of immunological alterations in psychiatric diseases has expanded the field; presently, immunological correlates of psychosomatic diseases and personality are sought. On the other hand, while immunological disease has been psychologically assessed for many years, a clear‐cut link between psyche and immunological changes has yet to be shown. This fact, along with the therapeutic implications of advancing knowledge, will influence strongly the future trends of psychoneuroimmunology. Concepts emerging from the study of this field will be of heuristic value to both psychiatry and immunology and will help define new and expanded limits for both disciplines.
Use of chernoff faces to follow trends in laboratory data
Authors: Lott John A. , Durbridge Timothy C.
Published: March 2007

Graphical representation of laboratory data speeds up recognition of abnormalities. The Chernoff face is an excellent tool to represent patient data; the human eye and mind are exquisitely sensitive to subtle changes in facial appearances and expression. One of the best uses of the faces is in illustrating trends in the data, particularly in acutely ill patients where many laboratory studies are performed. The faces are especially useful if comparisions to a former or normal state are made. A very brief examination of adjacent faces suffices to identify changes in laboratory data.