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7-(3-Amino-1-propynyl)-1-cyclopropyl-6,8-difluoro-1,4-dihydro-4- oxoquinoline-3-carboxylic acid (7a) and some related compounds (7b-f, 8a, b, 9) were prepared via palladium(0)-catalyzed cross-coupling reaction of 7-iodoquinolone 12 with acetylenic compounds and their antibacterial activity was tested. The methylene homologue (7d) and the N-methyl derivative (7e) of 7a showed essentially the same activity as that of 7a. Addition of methyl group(s) to C'-3 of 7a (giving 7b, c) reduced the activity. The hydrogenation of 7a to (Z)-3-amino-1-propenyl (8a), (E)-3-amino-1-propenyl (8b) and 3-amino-1-propyl (9) compounds retained or enhanced the activity of 7a. Among the compounds prepared, 8a was the most active, but was less active than ciprofloxacin (1). In order to get insight into structure-activity relationships, the spatial distribution of the amino groups of 7a, 8a, b, and 9 was examined by means of computer-aided molecular modeling.

A 44-year-old man with pemphigus vegetans had severe odynophagia. He received an endoscopic examination for esophageal involvement. Many white plaque-like lesions with an erythematous base were seen on the esophageal mucosa. Biopsy from the mucosal epithelial layer showed rounded epidermal cells with large nuclei and numerous inflammatory cells including eosinophils.

We presented an unusual case of cutaneous and bilateral ovarian metastases of malignant melanoma. There was no previously identifiable cutaneous or mucous lesion or teratoid element. Past history revealed that the patient had undergone removal of a blackish, elevated, irregular shaped tumor from the right arm eleven years previously. This lesion was considered to be the primary melanoma; therefore, it was thought to be very unusual as metastasis occurred in both the skin and ovaries after an eleven year disease-free interval.

Melanoma among Japanese is rare, and differs in its clinical and histological characteristics from that found in Caucasians. In this study, the ganglioside expression of melanoma specimens obtained from Japanese was determined and compared with previously published data on Caucasians. The ganglioside composition of 25 biopsy melanoma specimens, including 13 primary and 12 metastatic lesions, was studied using thin layer chromatography. Four gangliosides (GM3, GD3, GM2, GD2) were most commonly expressed in melanomas in Japanese. The expression of gangliosides was quite variable in both primary and metastatic melanomas as seen in previous reports. No significant differences were observed between gangliosides from primary and metastatic sites. A new type of ganglioside expression, in which GM3 was nearly the only ganglioside (> 95%), was found in metastatic tumors from two Japanese patients with acral lentiginous melanoma (ALM), which is the most common clinical and histopathological type of melanoma among Japanese but is very unusual among Caucasians. The patterns of expression were similar to those in Caucasians except for the detection of a "new" pattern.

In order to elucidate the mechanism(s) behind the interactions between barbiturates and Ca2+ antagonists, the effects of pentobarbital combined with three structurally diverse types of Ca2+ antagonist on CaCl2-induced contractile responses of the guinea pig thoracic aorta in Ca(2+)-free and 40 mM K+ medium and the effects of pentobarbital on Ca2+ antagonist binding to guinea pig aortic membranes were investigated. The dihydropyridine derivatives isradipine (10(-10)-10(-8) M) and nifedipine (10(-10)-10(-8) M) inhibited CaCl2-induced contractions concentration-dependently. Treatment with both pentobarbital (10(-4) M) and dihydropyridine Ca2+ antagonists (10(-9) M) shifted the CaCl2 concentration-response curves to the right significantly compared with those after treatment with the Ca2+ antagonists and pentobarbital alone. However, no synergistic effects of pentobarbital (10(-4) M) with other types of Ca2+ antagonist (verapamil (10(-7) M) and diltiazem (10(-6) M)) were observed. The binding of [3H]isradipine (2 x 10(-9) M) to guinea pig aortic membranes was increased significantly by simultaneous pentobarbital treatment, but no such effect was observed with [3H]verapamil (10(-8) M) or [3H]diltiazem (2 x 10(-8) M). These findings suggest that the synergistic contractile effects of pentobarbital and dihydropyridines were, in part, due to enhancement of dihydropyridine binding to guinea pig aortic membranes (L-type Ca2+ channels) by pentobarbital and that the interactions between pentobarbital and Ca2+ antagonists may be structurally specific.

We have selected GD3-deficient human melanoma cell lines, in order to investigate the function of GD3 ganglioside. This was done by treating SK-MEL-28 cells with anti-GD3 antibody (R24) and rabbit complement and subsequent subcloning of the surviving cells, resulting in the derivation of two cell lines deficient in the cell surface expression of GD3. Neither cell line (designated SK-MEL-28-N1 and SK-MEL-28-N2) had detectable cell surface expression of GD3 as analyzed with monoclonal antibody R24, and no GD3 was detectable in either cell line by glycolipid isolation, thin-layer chromatography, or resorcinol-HC1 spray, but thin-layer chromatography immunostaining with monoclonal antibody R24 showed the presence of low amounts of GD3 in both N1 and N2 (1/40 of the amount in the parent cell line in N1 and 1/500 in N2). In SK-MEL-28-N1, the residual GD3 was shown by immunofluorescence assays on permeabilized cells to be present in discrete intracellular organelles, suggesting that these cells have a defect in the transport of GD3 as well as in its synthesis. Both SK-MEL-28-N1 and -N2 had an increase in detectable GM3 expression. The mutant cell lines had altered cell morphology in comparison to the parent cell line and both had slower growth rates in vitro and lower tumorgenicity in nu/nu mice. These results indicate that GD3 ganglioside plays an important role in proliferation and growth of melanoma cells.

4-Substituted 6-cyclopropyl-6, 9-dihydro-5-methoxy-9-oxo-1H-imidazo[4,5-f] quinoline-8-carboxylic acids (6) and 8-substituted 1,5,6,11-tetrahydro-5-methyl-1-oxo-imidazo[4,5-g]pyrido[1,2,3-de][1,4] benzoxazine-2-carboxylic acids (7) were prepared as potential antibacterial quinolone derivatives. The appendages at C-4 of -6 and at C-8 of -7 were selected from 1-piperazinyl, 4-methylpiperazinyl, 3-aminomethylpyrrolidinyl, and 3-aminomethylpyrrolidinyl groups. The 5-methoxyimidazoquinolones 6 were superior to the corresponding ofloxacin type analogues 7 in in vitro antibacterial activity. The activity of 6 was equipotent against S. aureus, but 2 to 16 times less potent against E. coli and P. aeruginosa compared to that of the 5-fluoro analogue 3.

4,5-Disubstituted 6-cyclopropyl-6,9-dihydro-9-oxo-1H-imidazo (30-32) and triazolo[4,5-f]quinoline-8-carboxylic acids (33-35) were synthesized starting from 5,6-diaminoquinolones 25. The imidazoquinolones 30-32 were equal or superior to the corresponding triazoloquinolone analogues 33-35 in in vitro antibacterial activity. As for the C-5 substituents, a fluorine atom was the most favorable of the three groups, H, F, and Cl. Among the compounds prepared, 4-(cyclic amino)-5-fluoro-imidazoquinolones 31 a-d showed potent and well-balanced antibacterial activity against both gram-positive and gram-negative bacteria. Structure-activity relationships for the C-4 substituents (cyclic amino groups) were also examined in detail.

Mouse monoclonal IgM antibody (MAb 202) can cause melanoma cell necrosis in vivo. We analysed its immune mechanism in three melanoma patients to whom MAb 202 was administered. After the MAb 202 administration, histopathological analysis showed necrosis of melanoma cells expressing only GM3 in two patients. Another patient carrying both GM3 and GD3 showed infiltration of lymphocytes within the tumor nest but no tumor cells or nest necrosis. Immunohistological examination using anti-mouse IgM antibody revealed MAb 202 bound on the surface of melanoma cells in two patients but not in the third (positive for both GM3 and GD3). In vitro, MAb 202 reacted with the melanoma cells of the same two patients, but not with any other tissues of these individuals. We found no reaction of MAb 202 to non-melanoma cells including normal melanocytes and glia cells. Our trials suggest, 1) MAb 202 reacts directly to monosialogangliosides on the melanoma cell surface and then leads to the cytotoxicity reaction, or 2) MAb 202 induces lymphocyte infiltration and possibly then promotes the secretion of some cytokines.

ETs-induced contractions were resistant to ET(A)-selective antagonists and believed to be mediated by activation of ETB receptors in guinea pig bronchus. In the present study, the effects of the ET antagonists, PD143296 (Ac-D-Phe-L-Leu-L-Phe-L-Ile-L-Ile-L-Trp.2Na) and PD145065 (Ac-[(R)-2-10, 11-dihydro-5H-dibenzo[a, d]cyclohepten-5-yl]Gly)-L-Leu-L-Asp-L-Ile-L-Ile- L-Trp.2Na), on contractions induced by ET-1, ET-3, sarafotoxin S6c (STXc), and IRL1620 in the isolated hilar bronchus of the guinea pig were investigated. An ETA/B nonselective antagonist, PD145065 antagonized contractions induced by ET-1, ET-3, STXc, and IRL1620. Its antagonistic activity against ET-1, with pKB of 5.77 +/- 0.02 (n = 16, 3-10 microM), was significantly lower than that against ET-3, with pKB of 6.18 +/- 0.02 (n = 12, 3-10 microM), STXc, with pKB of 5.97 +/- 0.01 (n = 14, 3-10 microM), and IRL1620, with pKB of 6.80 +/- 0.04 (n = 14, 0.3-1 microM). Conversely, although a putative ETB-selective antagonist, PD143296 (10 microM) slightly but significantly antagonized the concentration-response curve of IRL1620 (pKB = 5.28 +/- 0.14, n = 6), it had no effect on ET-1-,ET-3-, or STXc-induced contractions. These results suggest that ETs possibly activate ETB2 or an atypical ETB receptor subtype in guinea pig hilar bronchus.

Characterization of the receptors mediating contractions to endothelin-1 (ET-1), endothelin-3 (ET-3), sarafotoxin S6c (STXc), or IRL 1620 in isolated epithelium-denuded hilar bronchus of guinea pig using as antagonists BQ-123 (ETA receptor-selective) and Ro 46-2005 (ETA/B nonselective) was investigated. ET-1, ET-3, STXc, and IRL 1620 produced only contraction, and their concentration-response curves were obtained at the same concentration range (10(-10)-10(-7) M). The potency order was the following: STXc = ET-3 = ET-1 > IRL 1620. BQ-123 (10(-5)M) had no marked effect on the contraction induced by ET-3 or STXc, whereas it attenuated the response induced by high concentration of ET-1 (3 x 10(-8)-10(-7)M). The contraction induced by IRL 1620 was antagonized by BQ-123 (3 x 10(-6)-10(-5)M). Ro 46-2005 (10(-5)M) failed to inhibit the responses to ET-1 and ET-3. Ro 46-2005 (10(-5)M) slightly, but significantly, shifted the concentration-response curve for STXc to the right (pKB = 4.94 +/- 0.10, n = 7), and the maximum response was potentiated to about 127%. The curve for IRL 1620 was shifted in parallel by Ro 46-2005 (3 x 10(-6)-10(-5)M) to the right (mean pKB = 6.35 +/- 0.09, n = 8). These results suggest that ETB receptors primarily mediate contraction to ET-1, ET-3, STXc, and IRL 1620, and the relative inhibitory activities of ET antagonists vary with the agonist used. However, ET-1 and ET-3 might also activate non-ETB receptor or unknown mechanisms.

Various azaazulene derivatives were synthesized and their antiallergic activity was examined. The structure-activity relationship among various derivatives modified by introducing substituents at the 1-,2-, or 3-position of the azaazulene ring was investigated. The inhibitory activities on allergic histamine release of the compounds bearing a 5-tetrazolyl group at the 3-position were more potent than those of the corresponding compounds with other groups (CN, COOH, and CHO). The compounds substituted with amino, azide and carboxymethylamino groups at the 2-position showed strong inhibitory activity. The compounds with various phenylalkyl groups at the 1-position showed a greater activity than those with other substituents. Among the compounds with substituents at the 1-,2-, or 3-position of the azaazulene ring, 1-benzyl-7-isopropyl-3-(5-tetrazolyl)-1-azaazulen-2-one (18f) and 1-(4-fluorobenzyl)-7-isopropyl-3-(5-tetrazolyl)-1-azaazulen- 2-one (19c) had the most potent inhibitory activities on histamine release from mast cells and on passive cutaneous anaphylaxis (PCA) in rats after oral administration (ED50 = 0.56 and 0.58 mg/kg, respectively).

A 45-year-old man with diffuse infiltrates on the chest X-ray film was admitted to the hospital. At first, sarcoidosis was suspected due to the elevated serum angiotensin-converting enzyme (ACE) activity. Transbronchial lung biopsy and bronchoalveolar lavage, however, did not lead to that diagnosis. Pulmonary alveolar proteinosis (PAP) was diagnosed after an open lung biopsy. The diffuse infiltrates on his chest X-ray film diminished spontaneously. Two years later, diffuse infiltrates increased again with an elevated serum ACE activity. In this case, it seemed that the severity of PAP was associated with serum ACE activity. We review the literature on the relationship between serum ACE activity and dysfunction of alveolar macrophages in PAP.

Endothelin is a potent bronchoconstrictor peptide first identified as a novel vasoconstrictor produced by vascular endothelial cells. Recent reports suggest that airway epithelial cells are also capable of releasing this active peptide. To investigate the regulatory mechanism of endothelin expression, we studied the effects of endotoxin and pro-inflammatory cytokines such as interleukin-1 and tumour necrosis factor on the expression and release of endothelin-1 by airway epithelial cells. Both endotoxin and the cytokines stimulated endothelin-1 release by human bronchial epithelial cells. Northern blot analysis showed increased expression of preproendothelin-1 mRNA by these factors. These results suggested that airway epithelial cells might play a role in the local airway smooth muscle tone through the production of endothelin, which might be upregulated by inflammatory products in the airways.

Hepatitis C virus (HCV) is a new virus discovered in 1989. Since HCV is known to cause fibrotic changes in the liver, we studied whether HCV is involved in the pathogenesis of idiopathic pulmonary fibrosis (IPF). Firstly, we assessed anti-HCV antibodies by enzyme-linked immunosorbent assay (ELISA) in the sera obtained from 66 IPF patients (46 males and 20 females; mean age +/- SEM, 61.5 +/- 10.1). We observed a significantly high prevalence of anti-HCV antibodies in IPF compared with 9,464 age-matched controls (28.8% vs 3.66%, p < 0.05). To confirm the results, recombinant immunoblotting assay (RIBA) was conducted on the 19 ELISA-positive sera, and 8 sera (12.2%) were found to be definitely positive. Secondly, we searched for HCV in the blood of IPF patients by reverse transcription-polymerase chain reaction. As preliminary data, four out of 28 cases (14.3%), all of which were pathologically diagnosed as UIP, were positive for HCV. In conclusion, although further investigation is required, a high prevalence of anti-HCV antibodies and the existence of HCV itself in the blood may suggest the possibility that HCV infection plays an important role in the pathogenesis of IPF.

A 65-year-old woman with facial erythema and hypergammaglobulinemia developed excessive fatigability. A diagnosis of Lambert-Eaton myasthenic syndrome (LEMS) was made from electrophysiological studies. She had symptoms and laboratory data compatible with probable Sjögren's syndrome. Skin biopsy revealed the histological findings of discoid lupus erythematosus. Treatment with 3,4-diaminopyridine resulted in the improvement of fatigability. LEMS should be recognized as a treatable complication of systemic autoimmune diseases.

Scatterplots of the Coulter STKS were studied in order to make better analyses of morphological abnormality of leukocytes. The analytical pattern of a sample failed to completely prevent blood coagulation, especially platelet aggregation, showed the poor separation between lymphocyte and neutrophil populations. On the scatterplot, a small population was occasionally observed in the lower area of normal lymphocyte population with elevation of room temperature, being thought an artificial population. When atypical and/or abnormal lymphocytes increased, the pattern was characterized by the distribution of large lymphoid cells spreading over monocyte population area. The characteristics became clear in proportion to the percentage of large lymphoid cells. To detect immature granulocytes, we introduced a criterion originated in our laboratory. Using the detection criterion, we could obtain the satisfactory results with sensitivity of 85%, specificity of 93%, efficiency of 91% and coefficient correlation (r) of 0.73. It is concluded that the pattern analysis and the detection criterion are useful in the experiments using routine laboratory samples and valuable in clinical implication.

We report cases of angina pectoris or minimal acute myocardial infarction accompanied by pulmonary edema, which were retrospectively studied with regard to their clinical characteristics, prognosis and treatment. Sixteen patients, 5 males and 11 females with a mean age of 72.6 years, admitted to the Cardiovascular Center of Sendai between January 1986 and June 1989, were studied. Ten had previous myocardial infarction. Hypertension, chronic renal failure and diabetes mellitus were found in 10, 7 and 7 patients, respectively. Electrocardiograms during cardiac ischemic attacks showed ST elevation in 8 and ST depression in the other 8 patients. Coronary arteriography which was performed in 6 patients revealed three-vessel disease in 5, and two-vessel disease in one. Mechanical ventilation was indicative of 7, and intraaortic balloon counterpulsation in 2 patients. Coronary artery bypass graft surgery was performed for 3 patients. All patients recovered from pulmonary edema and were discharged. During the mean 15-month-follow-up period, 8 patients died. The causes of death were sudden cardiac death in 3, acute myocardial infarction in one, congestive heart failure in one, post-surgical death in one, and non-cardiac death in 2.

A series of C-7 thio-substituted 1-cyclopropyl-1,4-dihydro-4-oxoquinoline-3-carboxylic acids were prepared and tested for their antibacterial activity. Structure-activity relationships associated with the C-5 and C-7 substituents were discussed. Among the C-7 substituents including alkylthio, arylthio, heteroarylthio, and cyclic aminothio groups, a 2-aminoethylthio group was the best for enhancing in vitro antibacterial activity. The C-5 variants increased activity in the order OH less than F less than H less than NH2. Of compounds prepared in this work, 5-amino-7-(2-aminoethyl)thio-1-cyclopropyl-6,8-difluoro-1,4-dihydro-4 -oxo-quinoline-3-carboxylic acid (18) was the most active.

We surveyed the clinical characteristics, treatment, and prognosis of 162 patients with unstable angina, who were admitted to our center between 1985 and 1987. There were 112 males and 50 females, with a mean age of 65 years. The clinical characteristics according to the American Heart Association classification were new angina of effort in 21%, changing pattern in 61%, and new angina at rest in 18% of the patients. ECG recordings during attacks of angina were obtained in 70%, and ST elevation was detected in 11%, ST depression in 54%, and T wave abnormality in 5%. Coronary arteriography performed in 42% of the patients revealed single vessel lesion in 21%, two vessel lesion in 10%, three vessel lesion in 5%, and left main trunk lesion in 3% of the patients. Seventy-seven percent of the patients were controlled by medical therapy, including nitrates, calcium antagonists, and, in some cases, beta blockades. Three percent of the patients were controlled with intra aortic balloon pumping in addition to medical therapy. Coronary artery bypass graft surgery (CABG) was performed in 6% of the patients. Since 1987, percutaneous transluminal angioplasty (PTCA) was introduced in our center and PTCA was performed in 9 patients (6%). Restenosis of the dilated portions of the coronary artery was observed and PTCA was again performed in 2 of 9 patients (22%). All patients who received CABG or PTCA survived and have been free from angina or myocardial infarction. Non-fatal myocardial infarction occurred in 10 cases (5.6%) and fatal infarction occurred in one patient (0.6%).

A poxvirus isolated from the respiratory tract of raccoons in a forest and swamp area near Aberdeen, Maryland, was characterized by biological, serological, and biophysical methods. The virus was shown to be related to the vaccinia-variola subgroup by serological and biophysical methods, but measurably different from the other viruses of this group by biological methods. It causes flaccid paralysis in 1-day-old suckling mice and does not grow well on CAM after two or three passages.

Serological surveillance of suspected orthopoxvirus infections in man is important for confirming the success of the worldwide smallpox eradication programme. An adsorption radioimmunoassay (RIA) was used to differentiate sera from patients who were naturally infected with human monkeypox or variola virus, and individuals who were immunized with vaccinia virus. The antisera were adsorbed with uninfected chicken chorioallantoic membrane (CAM) and vaccinia-infected CAM before reacting in RIA with vaccinia, monkeypox, and variola antigens. Each serum group showed characteristic patterns of residual antibody activity which made it possible to identify antibody specificities.When 45 human sera were tested by this method, 71% were identified as having vaccinia, variola, or monkeypox adsorption characteristics, while the remaining 29% could not be identified. Of the identified sera, 9 were characteristic of vaccinia, 8 of variola, and 15 of monkeypox. Six of the 15 monkeypox sera were virologically confirmed monkeypox infections, 6 were suspected monkeypox infections but were not virologically confirmed, and 3 were of unknown aetiology.The adsorption RIA provides a method of identifying serologically the poxvirus responsible for infection long after the acute phase of illness.

A 21-year-old Indian who worked as a vaccinator in the smallpox programme developed an atypical case of variola major despite several previous, successful vaccinations. The clinical course of the disease was unusual, as there were two distinctly separate "crops" of smallpox. The disease began less than one month after a successful vaccination, which therefore gave the briefest period of protection recorded for this reliable immunization. The patient was found to have a virtually complete IgM deficiency with normal levels of circulating antibody against orthopox virus.

Immunodeficient patients who were presumed to be susceptible received zoster immune globulin prophylaxis after exposure to varicella. The highest clinical attack rate (35.9%) was seen in household contacts; the lowest attack rate (0%) was observed in children exposed at school. Among household contacts, 48 of 100 patients who received high titer ZIG (reciprocal complement fixation titer greater than or equal to 2,560) developed fourfold rises in serum CF antibody between pre- and 48-hour post-treatment specimens, compared to only one of 34 patients treated with lower titer ZIG lots (P less than 0.001). Patients who developed fourfold antibody rises were significantly less likely to contract clinical varicella (P less than 0.01). Patients who received high titer ZIG also had significantly lower risks of death (P = 0.025) and complications (P = 0.006). Among ZIG-treated patients who contracted clinical varicella, 80% developed mild disease (less than 100 pox), and the median incubation period was prolonged. Immunodeficient children exposed to varicella benefit from ZIG prophylaxis and higher titer ZIG is of greatest benefit.

Although hospital-associated spread of vaccinia has been reported in the past, there have been no recent reports. This paper describes hospital-associated spread of vaccinia virus infection, supplies data on the environmental survival of vaccinia virus and offers recommendations for the management of patients with vaccinia that may minimize the hazard of infection in other high-risk patients.

Epidemiological studies of measles and measles immunization frequently require determination of measles antibody status. In developing countries, where venipuncture is frequently unacceptable and where refrigerated storage of serum specimens is often unavailable, microtiter techniques not requiring refrigeration are required. We developed a filter paper technique that measures measles hemagglutination inhibition antibody and meets these criteria. Comparison of separately collected venous blood and peripheral blood collected on filter paper demonstrated 97% agreement in terms of presence or absence of antibody. In 30 of 32 measles specimens, 94% of titers were the same or varied by less than 2 twofold dilutions.

The specificities of antisera during development of the humoral antibody response to poxvirus antigens were examined in monkeys injected with chimp-9 whitepox virus or monkeypox virus. Sera were obtained from 3 African green (vervet) monkeys inoculated with chimp-9 whitepox virus, 1 rhesus monkey inoculated with monkeypox virus, and 2 rhesus monkeys inoculated with soluble monkeypox viral antigen. The sequentially obtained sera from each animal were adsorbed with uninfected chicken chorioallantoic membranes (CAM) or vaccinia virus-infected CAM. The adsorbed sera were tested by radioimmunoassay to determine the specificity of the residual antibodies to vaccinia, variola, and monkeypox viruses. The adsorbed sera at different stages of the immune response showed increasing specificity with time after inoculation. Generally, antibodies in sera collected earlier than 21-27 days after immunization could not be identified after adsorption, but late sera could be identified unequivocally.

The results are presented of a special study to determine whether variola-like "whitepox" viruses could arise as white pock variants of monkeypox virus after one or a few mutations. DNA mapping by cross-hybridization of restriction endonuclease DNA fragments was carried out on 18 orthopoxviruses relevant to this study, including variola and monkeypox viruses and white (non-haemorrhagic) pock producers recovered from chorioallantoic membranes infected with red (haemorrhagic) pock-producing monkeypox viruses. The distinctiveness of the DNA maps of true variola and monkeypox viruses indicated that spontaneous production of "whitepox" from monkeypox virus was genetically impossible. These and other observations led to the conclusion that the "whitepox" viruses recovered from monkeypox virus stocks had an exogenous origin.

Monkey pox virus was mechanically disrupted by low temperature and high pressure into soluble and insoluble fractions. Soluble fractions elicited virus-neutralizing antibodies (1:20 to 1:160) in rabbits, whereas the insoluble (in saline) fractions did not (less than 1:5). No infectious virus was detected after the disruption procedure. Rhesus monkeys immunized with the soluble fraction elicited virus-neutralizing (1:1,200), complement-fixing (1:16), and hemagglutinating-inhibiting (1:80 to 1:160) antibody titers and were completely protected against monkey pox virus-induced disease. This model of monkey pox virus subunit vaccine preparation may prove to be useful in developing an efficacious noninfectious vaccinia vaccine for use in high-risk individuals.

An enzyme-linked sandwich immunoassay using human thyroglobulin conjugated with beta-D-galactosidase and silicone rods coated with human thyroglobulin was developed for the measurement of circulating anti-thyroglobulin autoantibody. The volume of serum needed for the assay was as little as 5 microliter. The sensitivity of the assay was approximately 7 x 10(-15) mol/tube of antithyroglobulin immunoglobulin G corresponding to 220 ng/ml of serum, which was equal to or rather higher than that of radioimmunoassay. The specificity of the assay was demonstrated by (1) parallelism of the standard curve with dilution of sample sera of patients with thyroid diseases, and (2) non-detectability of anti-thyroglobulin in autoantibody in the sera of normal subjects. The precision of the assay was proven by (1) sufficient recovery of antithyroglobulin immunoglobulin G added to serum, and (2) coefficients of variance within and between assays were 6.5 to 10.3% and 4.9 to 14.1%, respectively. No effect of thyroglobulin on the present assay was observed when the ratio of the amount of thyroglobulin to that of anti-thyroglobulin immunoglobulin G was lower than 1 : 10. Furthermore, a significant correlation was observed between anti-thyroglobulin autoantibody concentrations measured by our enzyme immunoassay and those by tanned red cell hemagglutination (r = 0.78), and between those by the enzyme immunoassay and those by radioimmunoassay (r = 0.80). The application to clinical samples ensured the high sensitivity and the adequate validity of the present assay.

The results of the Grice extra-articular subtalar arthrodesis were evaluated in 102 feet of 60 ambulatory patients with spasticity at an average of five years postoperatively. Results were satisfactory in 96 feet (94 per cent). Unsatisfactory results were attributed to recurrent deformity in four feet and over-correction in two. No other significant complications were identified. The Grice arthrodesis is recommended for hindfoot valgus deformity secondary to spasticity which is refractory to non-operative management.

In September, 1987, a survey on the nutrition of infants was conducted with 1,235 children from 3 to 6 years of age selected from six nurseries and six kindergartens in the metropolis of Tokyo. We compared the survey results with other similar survey reports with respect to three areas; 1, caries attack on primary teeth, 2, anomalies of dentition and occlusion and 3, how foods are eaten, to observe the difference by age and also between children in nurseries and in kindergartens. The following are the results of this comparative investigation. 1) The ratio of caries attacks on the 4-year-old in this survey was 69.6%, which was lower than the 83.4% of the nationwide survey of 1987 conducted by the Ministry of Health and Welfare, while the average value of the df index per person was 4.7, which was lower than the 6.1 of 1981. 2) The caries incidence rate was higher with children in kindergarten than those in nurseries and many kindergarten children had had more than one caries attack. 3) The frequency rate of anomalies of dentition and occlusion in the entire survey sample was 12.9% for deep overbite, 11.4% for open bite and 10.7% for edge-to-edge bite. 4) In the survey of how foods were eaten, the question, "Does your child like to eat things that are hard to bite on?" was asked, and 10.4% of the group answered "My child prefers eating soft food," which was quite high. To the question, "Does your child often eat things that are difficult to eat?", 17.1% of the entire group answered, "My child refuses to." This tendency was greater among the children in kindergaretens.

To differentiate antibodies to monkeypox and vaccinia viruses, the solid-phase ELISA with preliminary adsorption of sera with vaccinia virus was used. The identification of antibodies in 99 serum samples obtained in the Republic of Congo from children without vaccination scars was carried out by means of this assay. No antibodies to monkeypox virus were revealed. In 62% of cases the presence of antibodies was due to vaccinia virus. In the sera of 32% of children under examination antibodies differing from those to vaccinia and monkeypox viruses were revealed. To find out the origin of these antibodies, further seroepidemiological studies are necessary.

An enzyme-linked sandwich immunoassay using silicone rods coated with rabbit (anti-human thyroglobulin) immunoglobulin G and rabbit (anti-human thyroglobulin) monovalent fragment of immunoglobulin F (Fab') conjugated with beta-D-galactosidase was developed for the measurement of thyroglobulin in human serum. The volume of serum needed for the assay was as little as 2 microliters. The sensitivity of the assay was 3.5 ng/ml, which is equal to or rather higher than that of radioimmunoassay. The specificity of the assay was demonstrated by the following observations: (1) The absence of crossreaction of thyroxine and triiodothyronine, (2) non-detectability of thyroglobulin in the sera of patients who underwent total thyroidectomy, (3) parallelism of the standard curve with dilutions of reference serum. The precision of the assay was proven by the demonstration of the sufficient recovery of human thyroglobulin added to sera (92--99%) and coefficients of variance in within and between assays were 6.2--9.3 and 2.5--5.3%, respectively. Furthermore, a highly significant correlation was observed between thyroglobulin concentrations measured by our enzyme immunoassay and those by radioimmunoassay (r = 0.99, p less than 0.001, n = 63). Human thyroglobulin in serum was detectable in 90% of 146 normal subjects, the concentration (mean +/- S.D.) being 13.3 +/- 10.3 ng/ml.