Liver microsome-mediated activation of proestrogens in the environment is reviewed here. Proestrogens such as methoxychlor, trans-stilbene, diphenyl, diphenylmethane, 2,2-diphenylpropane, benzo[a]pyrene, benzophenone, 2-nitrofluorene (NF), chalcone, trans-4-phenyl-3-buten-2-one and styrene oligomers are negative in in vitro estrogen screening tests. However, those proestrogens exhibit estrogenic activity after metabolic activation by the microsomal cytochrome P450 system. In these cases, hydroxylated derivatives of the compounds are formed as major metabolites, and these metabolites exhibit significant estrogenic activities. Thus, the estrogenic activities of proestrogenic compounds are a consequence of metabolism of the parent compounds. Various candidates for proestrogens among medicines and insecticides are also discussed.

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Bone loss with aging induces osteoporosis. The most dramatic expression of the disease is represented by fractures of the proximal femur. Pharmacologic and nutritional factors may play a role in the prevention of bone loss with aging. β-Cryptoxanthin, a kind of carotenoid, is abundant in Satsuma mandarin orange (Citrus unshiu MARC.). Amoung various carotenoids including β-cryptoxanthin, lutein, lycopene, β-carotene, astaxanthin, and rutin, β-cryptoxanthin has been found to have a unique anabolic effect on bone calcification in vitro. Hesperidin, which is contained in Satsuma mandarin orange, did not have an anabolic effect on bone calcification in vitro. β-Cryptoxanthin has stimulatory effects on osteoblastic bone formation and inhibitory effects on osteoclastic bone resorption in vitro, thereby increasing bone mass. β-Cryptoxanthin has an effect on the gene expression of various proteins which are related to osteoblastic bone formation andmineralization in vitro. β-Cryptoxanthin has inhibitory effects on enzyme activity which is related to osteoclastic bone resororption, and the carotenoid induces apoptosis of mature osteoclastic cells in vitro. Oral administration of β-cryptoxanthin has been shown to have the anabolic effects on bone components in young and aged rats, and the administration has the preventive effects on bone loss in streptozotocin-diabetic rats and ovariectomized rats in vivo. Moreover, the intake of β-cryptoxanthin-reinforced juice for longer periods has been shown to have both stimulatory effects on bone formation and inhibitory effects on bone resorption in healthy human or postmenopausal women in evaluating with serum biochemical markers of bone metabolism in vivo. Thus the intake of dietary β-cryptoxanthin may have a preventive effect on osteoporosis due to stimulating bone formation and due to inhibiting bone resorption. Moreover, epidemiological studies suggest the potential role of β-cryptoxanthin as a sustainable nutritional approach to improving bone health of human subjects. β-Cryptoxanthin is an important food factor in maintaining bone healthy and in preventing osteoporosis.

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Nowadays, research on medicinal plants has attracted a lot of attention globally. A number of evidence has been accumulated to demonstrate promising potential of medicinal plants used in various traditional, complementary, and alternative systems. In recent years, a medicinal plant, Andrographis paniculata, and its major active phytochemicals have been extensively studied for several pharmacological activities. To understand the mechanism of action, researches have to be carried out at molecular levels. The present review aims at compiling consequential compendium of pharmacological benefits of health on this plant and its major diterpenoid constituent andrographolide that have been tested in various experimental models using modern scientific methodologies.

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To evaluate the developmental toxicity of decabromodiphenylether (DBDE) after exposure during the period from late gestation to after lactation, maternal Sprague-Dawley rats were given DBDE at dietary concentrations of 0, 10, 100, and 1000 ppm from gestational day 10 (GD 10) to postnatal day (PND) 21. On PND 21 and 77, lymphocytes (Lymph) in the spleen, thymus, and peripheral blood of male pups were subjected to flow cytometric analyses for expression of surface markers [CD3, CD4, CD8a, CD25, CD45RA, CD71, and CD161(NKRP1A)]. On PND 21, the proportions of splenic CD4+ T cells in the 10-ppm group, activated B (CD45RA+CD71+) cells in the 100- to 1000-ppm groups, and activated T cells (CD3+CD71+) in the 1000-ppm group were significantly decreased, and the population of peripheral CD161+ natural kiler cells on PND 21 and 77 had decreased in the 100- to 1000-ppm groups. In the 1000-ppm group, the serum T3 level was significantly decreased on PND 21 and the serum T4 level was decreased on PND 77. The body, spleen, and thymus weights were not significantly decreased, but liver weight was significantly increased on PND 21 in the 10- to 1000-ppm groups. These results suggest that on PND 21, developmental exposure to the highest dose of DBDE had a weak immunomodulatory effect. Although the most of the immunomodulatory effect had recovered to normal levels on PND 77, a decreasing effect on the natural killer (NK) cell population remained.

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Inorganic agents, such as Ag, Cu, and Zn compounds, are known to be relatively safe, and these agents are used in many products. In Europe, the standard levels of heavy metals eluted with artificial sweat and saliva are specified in the self-imposed safety criteria (OEKOTEX Standard) for textile products. We prepared metal zeolites (Ag, Cu, Zn, and Cr) and standard cloths loaded with these zeolites and a silver antimicrobial agent, AG300. The agents, standard processed cloths, and regions of commercially available products in which metals were detected at a high concentration were subjected to metal elution with artificial sweat (JIS L 0848: 2004), saliva (BS 6684: 1987), and purified water according to the JIS shake-flask antimicrobial test method (JIS L 1902-1900). The metal concentrations in the extracts were measured using an inductively coupled plasma-mass spectrometry (ICP-MS). A similar tendency was noted in elution from the agents, standard processed cloths, and commercial products. No metals were eluted with purified water alone, while the metals were eluted with artificial sweat and saliva. Large amounts of Cu and Zn were eluted, the elution of Ag was low, and almost no Cr was eluted. Furthermore, the antimicrobial activity of the standard processed cloths was evaluated in the antimicrobial test (JIS L 1902: 2002) using Staphylococcus aureus (S. aureus) and Klebsiella pneumoniae (K. pneumoniae). Cu- and Ag (Ag zeolite and AG300)-processed cloths exhibited strong antimicrobial activities against both bacteria. Zn-processed cloth also showed antimicrobial activity against S. aureus.

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Environmental pollution with polychlorinated biphenyls (PCBs) continues in Japan due to their high persistency, although their manufacture and use have already been prohibited. In this study, PCBs in sediments at 67 sites in Nagasaki Prefecture were determined by GC-MS, and their profiles were compared with those of commercial PCBs (Kanechlor KC300-600). The concentrations of PCBs in the sediments of sea and river areas in Nagasaki Prefecture were found to be up to 2.5 and 0.035 μg/g, respectively. Higher concentrations of PCBs in the sediments of the sea compared to those from river were observed (except area D vs N). The profile of PCB homologues in the sediments of the sea areas was similar to that of Kanechlor KC600, whereas the profile of PCB homologues in the river sediments was similar to that of Kanechlor KC500. Moreover, the source contribution was estimated by a chemical mass balance (CMB) method using the composition ratio of major congeners of Kanechlor. As a result, it was found that the contributions of KC600 and KC500 were higher in sea sediments (ca. 60%) than in river sediments (ca 50%).

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Cholinergic deficits are found in both vascular dementia and Alzheimer's disease (AD). Cholinesterase inhibition (CHI) is the only strategy that has been proven to have beneficial effects in patients, but may cause a broad spectrum of adverse events such as nausea, vomiting, and diarrhea, etc. To investigate how to attenuate the peripheral side effects of CHI agents without affecting their efficacy, the effects of propantheline bromide (PB) co-administered with donepezil on the gastric emptying (GE), gastrointestinal transit (GIT), brain cholinesterase (ChE) activities, and maze tasks of mice with memory impairment induced by transient ischemia were observed. The results indicated that PB decreased the increase in GE and GIT, but did not change brain acetylcholinesterase (AChE) activity or the latency of escape in cerebral ischemic mice treated with donepezil. These findings suggest that PB is nearly unable to penetrate the blood-brain barrier and could attenuate the peripheral side effects of CHI agents in the peripheral nervous system and without affecting their therapeutic effects in the central nervous system.

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Immediate-type hypersensitivity is involved in many allergic diseases such as asthma, allergic rhinitis, and sinusitis. The discovery of drugs for the treatment of allergic disease is an important subject in human health. Stimulation of mast cells releases inflammatory mediators, such as histamine, and proinflammatory cytokines with immune regulatory properties. We investigated the effect of the aqueous extract of Mosla dianthera (M. dianthera) (Maxim) (AEMD) on the immediate-type allergic reaction and studied its possible mechanisms of action using the model of mast cell-mediated allergic reaction. AEMD dose dependently inhibited compound 48/80-induced systemic allergic reaction and serum histamine release in mice. AEMD attenuated immunoglobulin E (IgE)-mediated skin allergic reaction and histamine release from mast cells. In addition, AEMD decreased the gene expression and production of tumor necrosis factor (TNF)-α in phorbol 12-myristate 13-acetate (PMA) and calcium ionophore A23187-stimulated human mast cells. Our findings provide evidence that AEMD inhibits the mast cell-derived allergic reaction and that TNF-α is involved in these effects. These findings indicate that AEMD could be a candidate as an antiallergic agent.

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Aqueous 4-methylphenol solution with bromide ion was treated with hypochlorite at 20°C under various experimental conditions. Changes in the composition of the halogenated products in water were determined by gas chromatographic (GC) and GC-mass spectrometric (GC/MS) analyses of diethyl ether extracts. 4-Methylphenol was shown to produce a variety of halogenated compounds, including polybrominated/chlorinated phenoxyphenols (Br/Cl-predioxins), having one to four halogen atoms, as by-products in the chlorine-treated water. Production of these predioxins is dependent on the number of equivalent of chlorine atoms per mole of compound and on the reaction pH. These results suggest that the chlorine treatment of water contaminated with 4-methylphenol and bromide ion leads to the formation of Br/Cl-predioxins, which are precursor of the highly toxic halogenated dibenzo-p-dioxins.

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To explore an optimal improvement of diabetic wound healing we have comparatively studied the effects of recombinant human acidic and basic fibroblast growth factors (rhaFGF and rhbFGF) on the healing impaired skin wounds in diabetic patients. Before using rhaFGF in diabetic patients, its pharmacokinetic features and possible toxic effects were evaluated using a rabbit model. For the 139 diabetic patients, 25% of them were divided into the group to receive topical rhbFGF daily at the dose of 100 U/0.1 ml/cm² as positive control (n=35), and the remainder to receive topical rhaFGF at the same dose as rhbFGF (n=104). Pharmacokinetic studies showed that plasma concentration of rhaFGF rapidly increased and reached to peak levels at 0.5 hr and then quickly decreased to normal levels at 3 hr after topical application on the wounds. No detectable toxic effects were found by examining multiple organs of the rabbits at 28 days after applying rhaFGF at 900 U/cm². Topical application of either rhbFGF or rhaFGF at 100 U/cm² effectively cured the skin wounds in the diabetic patients. Although there was no remarkable difference between groups, rhaFGF provided a slightly better healing rate and efficiency than rhbFGF did. Therefore rhaFGF will become an alternative candidate for diabetic wounds, especially when diabetic wounds were in acidic environment due to bacterial infection.

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The binding of advanced glycation endproducts (AGEs) to a cell-surface receptor for AGEs (RAGE) induces the formation of reactive oxygen species (ROS), which have been causally implicated in the pathogenesis of diabetic vascular complications. Pomegranate fruit extract (PFE) contains, a naturally occurring polyphenolic compound reported to possess potent radical-scavenging and antioxidant properties and to display significant cardiovascular protective action. In this study, we investigated whether PFE could inhibit glycated protein-iron chelate-induced toxicity by interfering with ROS generation in human umbilical-vein endothelial cells (HUVEC). Glycated fetal bovine serum (GFBS) was prepared by incubating fetal bovine serum (FBS) with high-concentration glucose. Stimulation of cultured HUVECs with 50 mM 1 ml of GFBS significantly enhanced lipid peroxidation and decreased antioxidant enzyme activities and levels of phase II enzymes. However, preincubation of the cultures with PFE resulted in a marked decrease in the level of lipid peroxide (LPO) and increase in the levels of antioxidant enzymes in a PFE concentration-dependent manner. These results demonstrate that PFE could inhibit LPO and enhance the antioxidant enzyme status in GFBS-iron chelate exposed endothelial cells by suppressing ROS generation, thereby limiting the effects of the AGE-RAGE interaction. Hence, PFE may have therapeutic potential in the prevention and treatment of vascular complications in diabetic patients.

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We have previously established a type of anti-resistant stealth liposomal topotecan plus amlodipine for overcoming the multi-drug resistance (MDR) in resistant leukemia cells. The objective of the present study was to further characterize it in the diversified non-resistant solid tumors in vitro and in vivo. Stealth liposomal topotecan plus amlodipine was re-prepared and physicochemically characterized. The in vitro drug release assays of topotecan and amlodipine from liposomes were performed using a dialysis method. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assays were performed in murine sarcoma S180 cells and human breast cancer MCF-7 cells, respectively. Apoptotic percentages of S180 cells were evaluated using flow cytometry. In vivo anti-tumor activity study and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) analysis were performed in male Institute of Cancer Research (ICR) mice with S180 xenografts. Stealth liposomal topotecan plus amlodipine exhibited high drug encapsulation efficiencies, suitable particle size distribution, negatively charged zeta potential and prolonged release profiles for both topotecan and amlodipine. Amlodipine potentiated the antiproliferative effect and inducing apoptotic effect of topotecan on the tumor cells, exhibiting an additive anti-tumor effect. Stealth liposomal topotecan plus amlodipine showed the optimal anti-tumor activity and inducing apoptotic effect in the in vivo studies. Stealth liposomal topotecan plus amlodipine demonstrated an overt anti-tumor activity in non-resistant solid tumors, suggesting that it deserves further clinical evaluations.

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Oxidative stress affects all intracellular macromolecules, and leads cells to death under unfavorable conditions. Glutathione (GSH) is known to play a critical role in the cellular defense against unregulated oxidative stress in mammalian cells including neurons. We previously demonstrated that GSH depletion induces cell death in the retina, but the mechanism of cell defense by GSH is still unclear. Thus, we here examined the effect of GSH depletion on expression of members of B-cell CLL/lymphoma 2 (Bcl-2) family (bcl-2, bcl-X_L, bax, bak, n-bak and bad) known to play key roles in determining cell viability. In order to deplete intracellular GSH, we systemically administrated buthionine sulphoximine (BSO), an inhibitor of γ-glutamylcysteine synthetase to mice. After 0, 1, 4, and 7 days of BSO injection, total RNAs from retina of each animals were isolated and subjected to real-time reverse transcription (RT)-polymerase chain reaction (PCR) analysis. Expression of bcl-X_L increased one day after BSO injection, but was back to the basal level on day 4. Its expression decreased on day 7. Expressions of bcl-2 and bax were significantly decreased from 4 days after BSO injection, whereas expression of bad was not changed. An anti-apoptotic molecule, bak displayed a significant decrease 7 days after BSO injection, whereas neuronal cell specific Bak (Bcl-2 homologous antagonist/killer), N-Bak was not altered in its gene expression. Taken together, we demonstrated that decrease in intracellular glutathione level altered expressions of Bcl-2 family members in distinct manners. Our study implies a defensive mechanism of GSH against oxidative stress for retinal neuronal survival which may involve alteration of expression of Bcl-2 family members, especially Bcl-2 and Bcl-X_L. Thus, over-expression of Bcl-2 and Bcl-X_L may not the only but a critical factor in GSH-dependent cellular protection, and which implies Bcl-2 and Bcl-X_L may provide a potent therapeutic tool for cures against oxidative stress induced retinal degenerative diseases such as glaucoma, retinopathy, and age-related macular degeneration.

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We measured the concentrations of acetone and isoprene in the exhaled breath from students to evaluate their lifestyles for the annual medical checkup at our university. 451 students were examined for both gases simultaneously by a gas analysis device, Biogas Acetone Analyzer (BAS-2000). The average concentrations of acetone and isoprene in the breath were 0.53±0.45 ppm and 0.065±0.058 ppm, respectively, and the frequency of the abnormal high levels by judging the cut-off values of 1.5 and 0.20 ppm were 3.5% and 2.7% of them, respectively. Acetone concentration in the breath was higher in students having no breakfast in the day of the medical checkup than those having breakfast. Furthermore, acetone concentration was significantly correlated inversely with body fat percentage in students who took the medical checkup in the morning and significantly higher in students with body mass index (BMI) under 25 than obese students with BMI over 25. On the other hand, isoprene concentration was higher in male students than female students, and also higher in smokers than nonsmokers. However, isoprene level was not affected by ingestion of breakfast in the day. Acetone and isoprene concentrations were correlated positively with each other, although the relationship was not affected by having breakfast. The results suggest that the measurement of acetone and isoprene concentrations in the breath is useful for evaluating lifestyles such as lack of breakfast, smoking and obesity in students.

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The object of this work was to investigate several adsorbents to determine their effective removal of benzene from wastewater. Tea and coffee lees were found to effectively adsorb benzene. Equilibrium adsorption isotherms conformed to the Freundlich isotherm (log-log linear). Adsorption of benzene by tea or coffee lees was observed in the pH range of 1 to 11. At equilibrium, the adsorption efficiency of tea or coffee lees for benzene was lower than that of activated carbon. The removal of benzene by tea or coffee lees was attributed to the uptake by intracellular particles called spherosomes. Ninety minutes after application of lees to wastewater (pH 10) containing 0.1 g/l of benzene, 75 to 82% of the benzene had been removed.

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Dyslipidemia appears to be a conventional risk factor for acute myocardial infarction (AMI). Several studies have reported decreased levels of antioxidants and increased levels of ischemia-modified albumin (IscMA) and lipid peroxides in dyslipidemic myocardial infarct patients. However, literature search reveals no reports of normolipidemic AMI patientswith reference to antioxidants and IscMA studies. Therefore, this study determined the endogenous levels of antioxidants and IscMA in normolipidemic AMI patients so that prospective measures could be taken to avoid acute coronary complications. The serum lipid profile, albumin, uric acid, total bilirubin, malondialdehyde, conjugated dienes and IscMA levels were determined in 165 normolipidemic AMI patients and 165 age/sex-matched controls. In addition, serum lipid concentrations were estimated by enzymatic methods. Endogenous antioxidants were significantly decreased in AMI patients compared with controls. In parallel with this, serum malondialdehyde (MDA) and conjugated dienes were significantly increased in AMI patients compared with controls. IscMA levels were significantly increased in AMI patients compared with controls. As for the serum lipid profile, total cholesterol (TC), cholesterol in low density lipoprotein (LDL) and triglycerides (TG) were higher in AMI subjects. High-density lipoprotein (HDL)-to-cholesterol and LDL-cholesterol-to-HDL-cholesterol ratios were also greater in AMI subjects. However, HDL-cholesterol was lower in AMI patients than control. AMI is a multifactorial disease that can arise even in normolipidemic subjects. The present study suggests that measuring of serum antioxidants and IscMA in normolipidemic patients would provide an index of oxidative stress and ischemia due to structural modifications of circulating albumin in serum.

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We investigated the release of formaldehyde (FA) upon the decomposition of 2-bromo-2-nitropropan-1,3-diol (Bronopol: BP), an FA-releasing substance widely used as an antimicrobial preservative in cosmetics and toiletries in many Western countries. The decomposition process producing FA mainly depends on temperature and pH. Notably, the pH of the BP-diluted buffer solution was the factor most influencing the release of FA. The release was markedly in higher alkaline buffer compared with acidic buffer. When a BP solution [0.1% (w/v)] prepared with a weakly alkaline buffer (pH 8.0) was stored at 25°C, the concentration of FA reached 30 ppm after 24 hr. On the other hand, under acidic conditions (pH 2.0), little FA was produced over 50 days. Homemade cosmetics (lotions and jells) were made [with 0.1% (w/v) BP] and stored at a constant temperature of 25°C for 30 days. The concentration of FA increased with time, reaching 20 ppm after 50 days. We investigated seasonal variation in the release of FA upon decomposition of BP in Osaka. BP test solution [0.1% (w/v)] was prepared with 0.005M sodium phosphate buffer (pH 6.0) and stored at ambient temperature for 30 days. The experiment was conducted three times in different seasons (mid winter, early spring, and early summer). It was found that the release of FA with decomposition of BP differed among the seasons.

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Proteolytic enzymes play a significant role in malignancy including, loss of growth regulation, invasiveness and metastases formation. Alpha-1 antitrypsin (A1AT) is a secretory glycoprotein produced mainly in liver and monocytes. It is the most abundant serine protease inhibitor in human plasma. Deficiency of A1AT is an inherited disorder characterized by reduced serum level of A1AT. Protease inhibitor Z (PiZ) and protease inhibitor S (PiS) are the most common deficient genotypes of A1AT. The association of deficient A1AT subtypes with several tumors such as primary liver carcinoma, lung cancer, bladder cancer and malignant hepatoma was reported. This study was aimed to test the relationship between A1AT genotypes Z and S and breast cancer in Jordanian female patients. Blood samples were collected from 111 patients. DNA was isolated and polymerase chain reaction (PCR) was performed to amplify the regions contain the Z and S mutations in exon V, and III, respectively. Genotyping of Z and S alleles was performed by restriction fragment length polymorphism technique using Taq1 restriction enzyme. Our results demonstrated that 100% of the breast cancer patients were homozygous for the normal allele Protease inhibitor MM (PiMM) and no PiZ and PiS genotypes were found. In conclusion, there is no relationship between A1AT deficient genotypes Z and S and breast cancer in Jordanian female patients.

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1α,25-Dihydroxyvitamin D₃ [1α,25(OH)₂D₃], the active form of vitamin D₃, is known to exhibit an anti-tumor properties and markedly suppresses the growth of various human cancer cells. We synthesized novel vitamin D analogs, 1α,25-dihydroxyvitamin D₃-26,23-lactam (DLAMs), having a lactam moiety in the side chain, and examined the effects on cell growth of human prostate cancer cells LNCaP. 1α,25(OH)₂D₃ significantly suppressed both the number of cells and cell viability. The mRNA expression of p21, well-known as a tumor suppressive gene, was clearly induced by treatment with 1α,25(OH)₂D₃ in LNCaP cells. The effects of 1α,25(OH)₂D₃ on the growth suppression of LNCaP cells was attenuated by the simultaneous addition of (23S,25S)-DLAM-1P. In a computer docking simulation, (23S,25S)-DLAM-1P bound to vitamin D receptor (VDR), and its lactam moiety may interfere VDR helix-12 folding. Its stereoisomer (23R,25R)-DLAM-1P did not influence cell growth regulated by 1α,25(OH)₂D₃. The expression of p21 mRNA induced by 1α,25(OH)₂D₃ was suppressed by (23S,25S)-DLAM-1P but not by (23R,25R)-DLAM-1P in LNCaP cells. In the absence of 1α,25(OH)₂D₃, neither (23S,25S)-DLAM-1P nor (23R,25R)-DLAM-1P regulated the cell growth of LNCaP cells. Thus, (23S,25S)-DLAM-1P interferes with the VDR signal and acts as a vitamin D₃ antagonist in cancer cells.

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Diesel exhaust (DE) is known to be one of the main causes of air pollution. Several studies have suggested that DE causes lung cancer, cardiovascular disease, abnormal reproductive function, and central nervous system damage as well as type I allergy in the airway. Type I allergy also plays a role in pathogenesis of endometriosis. In the present study, we examined the effect of exposure to DE on a rat model of endometriosis. Endometriosis was induced by autotransplantation of endometrium to the peritoneum in female Sprague-Dawley rats exposed to DE during prenatal and postnatal periods. Endometriotic lesions, normal peritoneum, and blood samples were obtained on days 4, 7, and 14 after autotransplantation. The extent of stromal proliferative lesions in the endometriosis model was greater in the rats of the DE exposure group than in those of the control group on day 14. Serum monocyte chemoattractant protein (MCP)-1 level was significantly higher in rats with endometriosis in the DE exposure group than in those in the control group on day 14. Results of this study suggest that DE exposure enhances the histologic and molecular pathology of endometriosis in rats.

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Synthetic vitamin K3 (VK3, 2-methyl-1,4-naphthoquinone, or menadione) has been shown to have anticancer activity in various human cancer cells. We developed an enzyme-linked immunosorbant assay (ELISA) for determination of VK3 in fetal calf serum (FCS) without any pretreatment. The monoclonal antibody against VK3, which was secreted from an established hybridoma cell line (3A3) has been proven to have highly specific binding to VK3 in cross-reactivity analysis. The full measuring range of the assay extends from 0.39 to 50 μg/ml of VK3. Based on validation analysis, this immunological analysis is a precise, accurate, and sensitive method for determination of VK3. This simple immunological method would deserve to be applicable clinically.

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