Biomedical subjects

H Miyakawa

Publications and source records attributed to H Miyakawa.

At least 19 recordsLinked to original sources

PubMed2006

Anti-p97/VCP antibodies: an autoantibody marker for a subset of primary biliary cirrhosis patients with milder disease?

We previously reported that 12.5% of primary biliary cirrhosis (PBC) sera reacted with a 95 kDa cytosol protein (p95c) that was subsequently identified as a p97/valosin-containing protein (VCP). The clinical features and course of the six anti-p97/VCP-positive PBC patients with Scheuer's stage 1 and 2 liver biopsies were monitored for an average of 15 years. This group was compared with 50 PBC patients that did not have detectable anti-VCP. Autoantibodies to a full-length recombinant p97/VCP were assayed by immunoprecipitation. All six PBC patients with anti-VCP had antibodies to the mitochondrial pyruvate dehydrogenase complex-E2 antigen as measured by an addressable laser bead immunoassay. The first was a male with no evidence of liver failure that died of cerebral infarction at the age of 85. The second was a 73-year-old female with Hashimoto's thyroiditis who has remained clinically stable without ursodeoxycolic acid (UDCA) treatment. Although the third had no HCV antibodies, he developed hepatocellular carcinoma at the age of 76 and died of renal failure at 78. The fourth was a 50-year-old female who remained clinically stable during follow-up and the fifth with Hashimoto's thyroiditis and stable liver function following UCDA treatment. The sixth was a male patient presenting a mild clinical course. The clinical course of these patients was in contrast to the 50 comparison group PBC patients who did not have anti-p97/VCP. As the six PBC patients with anti-p97/VCP antibodies had slowly progressive liver disease and no mortality related to autoimmune liver disease, our observations suggest that this autoantibody might be an indicator of a favourable prognosis.

Adenosine Triphosphatases↗

PubMed2006

Effects of high peak airway pressure on the expression of heat shock protein 70 in rat lungs: a preliminary study.

BACKGROUND: Heat shock protein 70 (HSP70) is induced by a wide variety of stresses in addition to hyperthermia. Recent studies have clarified that mechanical stretching and pressure overload can induce HSP70 in some tissues and cells. However, it remains unclear whether HSP70 is induced in stretch-subjected lungs, such as those under mechanical ventilation. This study was designed to investigate the effects of high peak airway pressure (PAP) ventilation on HSP70 expression in intact rat lungs. METHODS: Male Sprague-Dawley rats were randomly allocated to one of three groups: non-ventilated (anesthesia alone) control group; PAP 15 cm H(2)O group (P15); and PAP 30 cm H(2)O group (P30). The rats in the PAP groups were subjected to pressure-controlled assisted ventilation at the appropriate PAP for 30 min. Rats were killed at 12, 24 and 48 h after ventilation or anesthesia alone, and the lungs were removed. The lung tissues were processed for immunohistochemical and Western blotting analyses of HSP70. RESULTS: Following 30 min of pressure-controlled assisted ventilation, HSP70 expression in the P30 group was significantly up-regulated in bronchiolar cells and subepithelial tissues at 12 h, and this up-regulation continued throughout the observation period. In contrast, there were no significant differences between the control and P15 groups, although the expression of HSP70 was higher in the P15 group than in the control group at all time points. CONCLUSIONS: HSP70 was induced by high PAP ventilation, but its specific role and induction mechanism remain unclear. Therefore, further investigations should be encouraged.

Airway Resistance↗

PubMed2005

Direct evidence for mutual interactions between perineuronal astrocytes and interneurons in the CA1 region of the rat hippocampus.

Recent studies have demonstrated that astrocytes express a variety of ion channels and neurotransmitter receptors and can modulate the activity of neurons. Since a single astrocyte makes tight contacts with many neighboring neuronal cells, they can provide efficient and wide modulation of neuronal networks. Here, we provide direct evidence for mutual interactions between perineuronal astrocytes and interneurons in the stratum radiatum of the rat hippocampus. Direct depolarization of a perineuronal astrocyte suppressed the excitatory postsynaptic currents in an adjacent interneuron and increased the paired-pulse ratio, indicating that perineuronal astrocytes have a suppressive effect on presynaptic elements. Moreover, perineuronal astrocyte activation modulated the directly induced firing pattern of the interneuron, with initial facilitation and subsequent suppression. Conversely, direct firing of the interneuron depolarized the membrane potential and reduced the input resistance of the perineuronal astrocyte. These results directly demonstrate the existence of bidirectional interactions between neurons and perineuronal astrocytes.

4-Aminopyridine↗

PubMed2004

Olprinone improves diaphragmatic contractility and fatigability during abdominal sepsis in a rat model.

BACKGROUND: Respiratory failure with diaphragmatic fatigability is common in patients suffering sepsis or septic shock. However, the development and progress of diaphragmatic fatigability remains poorly understood, and no method has been established to treat fatigability. In this study, we hypothesize that neutrophil activation contributes to the development of diaphragmatic fatigability. We also sought to investigate whether a phosphodiesterase inhibitor, olprinone, improves diaphragmatic fatigability associated with abdominal sepsis and inhibits an increase in myeloperoxidase activity in diaphragmatic muscle. METHODS: Male Wistar rats were randomly assigned to a sham group, coecal legation perforation group (CLP), and a phosphodiesterase inhibitor (PDE) pretreated group. At 16 h after surgical procedure, the left hemidiaphragm was removed for the measurement of diaphragmatic contractility and fatigability. In addition, for the measurement of serial changes in myeloperoxidase activity, the right hemidiaphragm was also removed at 4, 8 or 16 h after the surgical procedure in each group. RESULTS: In a septic model involving rats, we observed that diaphragmatic muscles were fatigable and myeloperoxidase activity increased. We also demonstrated that intraperitoneal administration of olprinone improves diaphragmatic fatigability and inhibits an increase in myeloperoxidase activity induced by abdominal sepsis. CONCLUSION: Olprinone represents a potential therapy for cases of respiratory failure with diaphragmatic fatigability resulting from inhibition of neutrophil activation.

Abdomen↗

PubMed2004

Autoantibodies from primary biliary cirrhosis patients with anti-p95c antibodies bind to recombinant p97/VCP and inhibit in vitro nuclear envelope assembly.

We have reported previously that p95c, a novel 95-kDa cytosolic protein, was the target of autoantibodies in sera of patients with autoimmune hepatic diseases. We studied 30 sera that were shown previously to immunoprecipitate a 95 kDa protein from [(35)S]-methionine-labelled HeLa lysates and had a specific precipitin band in immunodiffusion. Thirteen sera were available to test the ability of p95c antibodies to inhibit nuclear envelope assembly in an in vitro assay in which confocal fluorescence microscopy was also used to identify the stages at which nuclear assembly was inhibited. The percentage inhibition of nuclear envelope assembly of the 13 sera ranged from 7% to 99% and nuclear envelope assembly and the swelling of nucleus was inhibited at several stages. The percentage inhibition of nuclear assembly was correlated with the titre of anti-p95c as determined by immunodiffusion. To confirm the identity of this autoantigen, we used a full-length cDNA of the p97/valosin-containing protein (VCP) to produce a radiolabelled recombinant protein that was then used in an immunoprecipitation (IP) assay. Our study demonstrated that 12 of the 13 (93%) human sera with antibodies to p95c immunoprecipitated recombinant p97/VCP. Because p95c and p97 have similar molecular masses and cell localization, and because the majority of sera bind recombinant p97/VCP and anti-p95c antibodies inhibit nuclear assembly, this is compelling evidence that p95c and p97/VCP are identical.

Adenosine Triphosphatases↗

PubMed2003

Optical detection of dendritic spike initiation in hippocampal CA1 pyramidal neurons.

Previous studies have shown that spikes can be generated in the dendrites of CA1 pyramidal neurons. Some have suggested that, in response to synaptic inputs, spikes are initiated near the soma and propagate back into the dendrites, but some recent studies have shown that intense synaptic inputs initiate spikes in the dendrite. Here, we report the optical detection of spike propagation along the apical dendrites of hippocampal pyramidal neurons. Rat hippocampal slices were stained with the fluorescent voltage-sensitive dye, JPW1114, and optical signals monitored using a 16 x 16 photodiode array system at a frame rate of 4 kHz. A stimulating electrode was placed at the boundary between the stratum (str.) lacnosum-moleculare and the str. radiatum to stimulate the Schaffer collateral, and fast and slow signal components were detected in the dendritic and somatic regions. By comparing the optical signals with whole-cell recordings, we confirmed that the fast component was due to a population of dendritic spikes in pyramidal neurons. The fast component appeared in dendritic locations near the input sites in response to synaptic activation, and signal onset at the soma was delayed by a few milliseconds compared with that at the input sites. Local perfusion of a Na(+) channel blocker near the soma eliminated the fast component at the soma, but had no effect on the fast component at the input sites. Our results indicate that dendritic spikes can be initiated in dendrites near the input site and propagate orthodromically toward the proximal dendrites and the soma.

Anesthetics, Local↗

PubMed2003

[Procedure and problem for short-term outcomes on off-pump coronary artery bypass grafting].

We performed off-pump coronary artery bypass grafting (CABG) in all cases without reoperation case from July, 2002. Advantage of off-pump CABG versus on-pump CABG which is reduced a number of perioperative complication and early patients recovery was previously demonstrated. In our institute, the mean number of grafts per patients was 4.7 +/- 1.3, and the rate of using arterial grafts was 99.5% in all cases without minimally invasive direct coronary artery bypass (MIDCAB). The mean hospital stay after operation was 10.8 +/- 2.8. It was shorted remarkably in comparison with on-pump CABG; 19.4 +/- 6. Furthermore, sever complication was not occurred in any cases after operation though high risk cases were increased. In the early cases, atrial fibrillation complicated frequently (32%), but using after magnesium sulfate it was remarkably decreased (8.4%). On the other-hand, attention is necessary for the infection caused by the increase of high risk patients. Therefore, we used vancomycin (VCM) at these cases from the viewpoint of prevention. Recently, we performed remnant omental transfer for the sever diabetes mellitus case which was used bilateral internal thoracic artery on CABG. It learned to get the early recovery which was necessary for the off-pump CABG by the above additional treatment.

Antibiotic Prophylaxis↗

PubMed2002

Optical monitoring of synaptic summation along the dendrites of CA1 pyramidal neurons.

The primary function of neurons is to integrate synaptic inputs and to transmit the results to other cells. Recent studies with somatic whole-cell recordings have shown that separate excitatory inputs to hippocampal or cortical pyramidal neurons are summated non-linearly. In the present study, we examined how postsynaptic potentials (PSPs) are summated along the dendrites employing fast optical voltage imaging techniques. Rat hippocampal slices were stained with a fluorescent voltage-sensitive dye (JPW1114) and optical signals were monitored with a 16 x 16 photodiode array system. Two independent input pathways were stimulated individually or in pairs through glass electrodes such that different locations of the dendrites received separate synaptic inputs. We found that (1) the summation of PSPs was sub-linear along the entirety of dendrites, (2) the blockade of GABA(A) receptors suppressed sub-linearity and (3) further blockade of GABA(B) receptors suppressed sub-linearity of the summation of separate inputs on apical dendrites. Our study demonstrates that pyramidal neurons integrate PSPs linearly along the entirety of dendrites; moreover, GABAergic inputs are responsible for maintaining sub-linear summation in CA1 pyramidal neurons.

Animals↗

PubMed2002

Effects of inducible nitric oxide synthase and xanthine oxidase inhibitors on SEB-induced interstitial pneumonia in mice.

The authors have previously reported that intratracheal instillation of staphylococcal enterotoxin-B (SEB) induced interstitial pneumonia (IP) in autoimmune-prone mice. SEB-reactive T-cells were critically involved in the development of IP in this model. Concern has arisen about the hazards of reactive oxygen species (ROS) and reactive nitrogen species (RNS) in the process of lung injury and fibrosis. Therefore, the involvement of nitric oxide (NO) and superoxide anion (O2-) in the pathogenesis of IP in this autoimmune-prone model has been investigated. Nitrite/nitrate levels were increased in bronchoalveolar lavage (BAL) fluid and serum from SEB-injected mice. The signal of the NO-(N-(dithiocarboxy) sarcosine)2-Fe2+ complex was detected in the SEB-injected lung and whole blood by electron paramagnetic resonance (EPR) spectroscopy. NO production was significantly decreased by aminoguanidine (AG) treatment. Xanthine oxidase (XO) activity in the lung, BAL fluid, and plasma was increased with instillation of SEB, and 4-amino-6-hydroxypyrazolo(3,4-d)-pyrimidine (AHPP) significantly inhibited XO activity. Moreover, both AG and AHPP significantly decreased production of pro-inflammatory cytokines, numbers of infiltrated cells in BAL fluid, and the area of thickened alveolar septa in the SEB-injected lung. In conclusion, the overproduction of nitric oxide and super oxide anion were implicated in the pathogenesis of interstitial pneumonia, and inducible nitric oxide synthase and xanthine oxidase inhibitors had protective effects against interstitial pneumonia in this model.

Animals↗

PubMed2002

Cellular responses to oral pathogens.

BACKGROUND: Several previous epidemiological studies, along with the results of more recent animal model approaches, have suggested a role for periodontitis in atherosclerosis. Such an association could be mediated by direct interactions of periodontopathic bacteria with host vascular tissues. METHODS: The interactions of Porphyromonas gingivalis with endothelial cells and macrophages in vitro were investigated relative to modification of low-density lipoproteins (LDL). RESULTS: P. gingivalis 381, its outer membrane vesicles, and the lipopolysaccharide (LPS) derived from these organisms were all shown to induce modification of LDL in the presence of the murine macrophage J774.A.1. Such alterations led to an increase in the migration of the particles through agarose gels. In addition, direct modification of LDL by strain 381 was demonstrated in the absence of macrophages. This latter property appears to be related to the potent protease activities of the bacterium. These properties may contribute to modification of LDL to forms which have been strongly implicated in cholesterol lipid accumulation in vascular tissues. P. gingivalis 381 also appears to induce cyclooxygenase-2 expression in endothelial cells as determined with human umbilical vascular endothelial cells (HUVEC). CONCLUSIONS: These in vitro results with vascular cells in culture suggest a molecular basis for a potential role for periodontopathic bacteria such as P. gingivalis in augmenting foam cell formation characteristic of atherosclerotic lesions.

Animals↗

PubMed2001

Low-threshold potassium channels and a low-threshold calcium channel regulate Ca2+ spike firing in the dendrites of cerebellar Purkinje neurons: a modeling study.

Various types of voltage-gated ion channels are distributed along the dendrites of neurons in the central nervous system. We have recently shown experimentally that the dendrites of cerebellar Purkinje neurons contain low-threshold voltage-gated Ca(2+) channels and low-threshold voltage-gated K+ channels. Although we found that these channels are involved in regulating the onset of Ca(2+)-dependent action potentials in the dendrites, we were unable to identify which of the known types of low-threshold Ca2+ channels and K+ channels were responsible, since there was no reliable method of discriminating between them. Here, we have built a detailed compartmental model of a Purkinje neuron by incorporating two types of low-threshold Ca2+ channel (T-type and class-E, or R-type) and two types of low-threshold K+ channel (A-type and D-type), in addition to another eight voltage-gated channel types, using a compartmental model neuron simulator. The model reproduces the basic features of the depolarization-induced responses of Purkinje neurons, such as fast Na+ spikes in the soma, Ca2+ spikes in the dendrites, the slow onset of Ca2+ spikes, repetitive Ca2+ spikes in the presence of TTX, the marked shortening of Ca2+ spike onset in the presence of 4-aminopydridine, and the longer Ca2+ spike onset in the presence of Ni2+. Our model shows that the D-type K+ channel and the class-E Ca2+ channel regulate the onset of depolarization-induced Ca2+ spikes in Purkinje neurons. These channels might be involved in integrating synaptic inputs in Purkinje neurons.

Action Potentials↗

PubMed2001

GABAergic control of synaptic summation in hippocampal CA1 pyramidal neurons.

The primary function of neurons is to integrate synaptic inputs and to transmit the results to other cells. It was shown previously that separate excitatory inputs to hippocampal pyramidal neurons are summated nonlinearly. In the hippocampus, responses of pyramidal neurons are influenced by GABAergic inputs in feed-forward or feedback manner, and also by oscillatory network activities. It is likely that these GABAergic inputs regulate the way synaptic inputs are summated. To examine the roles of GABAergic inputs on synaptic summation, we made whole-cell recordings from the cell bodies of CA1 pyramidal neurons in rat hippocampal slices while stimulating two independent input pathways with short interstimulus intervals, and examined the manner by which postsynaptic potentials were summated. We found that: 1) the summation of the perforant pathway and the Schaffer collateral pathway inputs was sublinear when the interval between two inputs was shorter than 30 ms, 2) the blockade of GABA(A) receptors partially suppressed the sublinearity, and 3) further blockade of GABA(B) receptors removed the sublinearity totally. We also found that 4) the summation was superlinear under the concomitant blockade of GABA(A) and GABA(B) receptors when the two inputs arrived with no delay. Thus our study demonstrates that GABAergic inputs are responsible for keeping the summation of two separate inputs on CA1 pyramidal neurons sublinear.

Animals↗

PubMed2001

Single nucleotide polymorphisms of the mannose-binding lectin are associated with susceptibility to primary biliary cirrhosis.

Although the immunopathogenesis of primary biliary cirrhosis (PBC) remains unknown, familial clustering of patients with PBC suggests an important role for genetic factors. In addition, recent data support the thesis that the mucosal immune response against intraluminal pathogens may be involved with the onset of PBC. Mannose-binding lectin (MBL) is a key factor in innate mucosal defenses and has several key single nucleotide polymorphisms (SNPs). To study whether MBL gene SNPs are associated with susceptibility to PBC, we studied 65 patients with PBC and 218 controls by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and sequence specific priming-polymerase chain reaction (SSP-PCR) to examine four polymorphic loci: two (H/L and X/Y) within the promoter region and the other two (P/Q and A/B) within exon-1. We also analyzed serum MBL concentrations. Interestingly, the prevalence of haplotype HYPA, leading to hyper-production of MBL, as well as HYPA/HYPA genotype were significantly increased in PBC compared to controls (0.53 vs. 0.44, P=0.031; 33.9%vs. 17.0%, P=0.003, respectively). Furthermore, individuals homozygous for HYPA had a significantly increased risk for PBC (odds ratio (OR)=2.51, 95% confidence interval (CI)=1.34-4.66). Our results demonstrate that the MBL genotype can be significantly associated with increased risk for PBC, and further, that increased production of MBL plays a critical role in immunopathogenesis.

Carrier Proteins↗

PubMed2001

Differences in anti-LKM-1 autoantibody immunoreactivity to CYP2D6 antigenic sites between hepatitis C virus-negative and -positive patients.

Anti-liver kidney microsome type 1 autoantibodies (anti-LKM-1) are known to be present in sera of autoimmune hepatitis type II and a subset of chronic hepatitis C patients. The autoantigen to anti-LKM-1 has been identified to be cytochrome P450 IID6 (CYP2D6) and the most frequently cited CYP2D6 antigenic sites of anti-LKM-1 in sera from autoimmune hepatitis type II patients spans the region aa 256-269. Other antigenic sites on CYP2D6 exist and have been identified in the two patient groups. However, most of these sites are concentrated on the carboxyl-terminal side of the protein, and the amino-terminal region has not been thoroughly investigated. Here, we have studied the antigenicity of the CYP2D6 amino region and compared reactivities between hepatitis C virus (HCV)-negative and -positive Japanese patient groups. A total of 34 anti-LKM-1-positive sera (eight with autoimmune hepatitis type II and 26 with chronic hepatitis C) were included. The immunoreactivity of patients' sera was examined against four conformational and one linear CYP2D6 peptide fragments. A defined antigenic site spanning aa 181-245 was found to react with 88% (7/8) of autoimmune hepatitis type II patients, as opposed to only 38% (10/26) of chronic hepatitis C patients. This was a significant difference (P< 0.043). Among these positively reacting samples, five of the seven autoimmune hepatitis type II sera and four of the ten chronic hepatitis C sera also reacted with a synthetic peptide spanning aa 256-269. Anti-LKM-1 thus may be able to recognize simultaneously at least two antigenic sites on the CYP2D6 protein, and reactivities against individual epitopes may differ according to HCV infectivity status.

Adult↗

PubMed2001

Neutrophilia and granulocyte colony-stimulating factor levels after cardiopulmonary bypass.

PURPOSE: The precise mechanism of neutrophilia after cardiac surgery is unknown. Granulocyte colony stimulating factor (G-CSF) can increase the number of leukocytes. The purpose of this study was to evaluate the relationship between serum G-CSF levels and peripheral blood leukocyte counts after cardiac surgery. METHODS: We prospectively studied 10 patients undergoing cardiac surgery (coronary artery bypass grafting) using cardiopulmonary bypass (CPB). Plasma G-CSF levels and neutrophil count were measured before induction of anaesthesia, at the end of surgery, and on the first postoperative day. These changes were compared with those in patients undergoing non-cardiac major surgery (control group). RESULTS: At the end of surgery, G-CSF levels increased (P < 0.01) in both groups, but were higher in the control than in the cardiac group (3,250 +/- 690 vs 194 +/- 29.5 pg ml(-1), respectively, mean +/- SEM, P < 0.01). On the first postoperative day, G-CSF levels were still high in both groups, and were still higher in the control (710 +/- 179 vs 122 +/- 19.9, respectively, P < 0.01). However, neutrophilia was greater in the cardiac group than in the control. G-CSF response correlated positively with neutrophilia in the control group (r = 0.656, P < 0.05) but not in the cardiac group. CONCLUSIONS: Our results indicate that changes in leukocyte count following cardiac surgery are unique to patients undergoing CPB. G-CSF plays an important role as the mediator of neutrophilia after non-cardiac surgery, but not after cardiac surgery with CPB.

Aged↗

PubMed2001

High sensitivity of a novel ELISA for anti-M2 in primary biliary cirrhosis.

The use of an ELISA for the detection of anti-M2, a specific autoantibody in primary biliary cirrhosis (PBC), has been common in Japan. However, there are some problems in the sensitivity of this ELISA, especially in PBC patients showing antimitochondrial antibody (AMA)-negative sera or low AMA titers by immunofluorescence. Recently, a new ELISA for anti-M2 was developed, using porcine heart mitochondrial protein as the antigen. We report here comparative studies of the new and the former anti-M2 ELISAs. Porcine heart mitochondrial protein was prepared and used as the antigen for the new ELISA for anti-M2. Sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of this protein showed three major M2 antigen proteins. As the second antibody, peroxidase-conjugated anti-human mouse monoclonal IgM, in addition to monoclonal IgG, was included. The sera of 171 PBC patients were examined. As controls, we examined the sera of 167 non-PBC patients and the sera of 115 normal controls. The cut-off index was set at 10 U/ml, based on the results for the normal controls. No sera from the non-PBC patients or the normal controls were positive for anti-M2 by either the new or the former ELISA. However, the positivity rate for anti-M2 in PBC patients with the new ELISA was 78%; in contrast, that with the former ELISA was only 54%; this difference was significant (P = 0.00001). In particular, in 65 patients showing AMA titers of 1:20 or less, the positivity rate with the new ELISA was 51%; in contrast, that with the former ELISA was only 17%. As the sensitivity of the new ELISA is significantly higher than that of the former ELISA, especially for sera from patients showing AMA-negativity or low titers of AMA, the new ELISA is considered to be more effective than the former ELISA for use in anti-M2 screening assays in patients with PBC.

Animals↗

PubMed2001

Tetra-butyl ammonium attenuates evoked release of acetylcholine from mouse hemidiaphragm preparation.

Tetra-butyl ammonium is a homologous member of mono-quaternary ammonium salts, and it has been reported to have a property of nondepolarizing neuromuscular relaxant. However, no neurochemical evidences exist that tetra-butyl ammonium may interfere with quantal release of acetylcholine from motor nerve terminals. In this study, using the neurochemical method, we investigated the effect of tetra-butyl ammonium on stimulation-evoked release of acetylcholine from mouse hemidiaphragm preparation. The preparation was loaded with [3H]choline (5 microCi/ml). Low concentrations of tetra-butyl ammonium (10(-5) M) had no effects. On the other hand, at concentrations of 4x10(-5) and 10(-4) M, this compound significantly reduced the [Ca2+]o-dependent release of acetylcholine from phrenic nerves. This finding indicates that tetra-butyl ammonium possesses a presynaptic inhibitory effect on acetylcholine release from the phrenic nerve terminal.

4-Aminopyridine↗

PubMed2001

False positive reaction in ELISA for IgM class anti-M2 antibody and its prevention.

Anti-M2 of anti-mitochondrial antibodies is recognized as the specific autoantibody detected in sera from patients with primary biliary cirrhosis (PBC). The IgG class and IgM class of this antibody can be separately measured using each ELISA. In the present study, false positive reactions were found in some sera from non-PBC patients such as acute hepatitis A, syphilis and rheumatoid arthritis using the IgM anti-M2 ELISA. They showed an increase of polyclonal IgM, and positivity for IgM anti-cardiolipin or rheumatoid factors, respectively. So, we developed a means to prevent these false positive reactions. First, dilutions of test sera at 1:1000-fold were carried out in addition to the original method at 1:100-fold. Secondly, some blocking reagents were added into the buffer system. By serum dilution, non-specific bindings disappeared in most samples other than showing an increase in polyclonal IgM. Moreover, the addition of suitable blocking reagents such as fetal bovine serum (FBS) and skimmed milk into the buffer system could prevent these non-specific bindings. From these findings, the procedure of optical serum dilution and the addition of suitable blocking reagents successfully prevented false positive reactions in this IgM anti-M2 ELISA.

Journal Article↗