Characterization of the soluble immune complex (EIC) of the amplified enzyme-linked immunosorbent assay (a-ELISA) and an evaluation of this assay for quantitation by reaction stoichiometry

The molecular composition of the soluble enzyme immune complex (EIC) of alkaline phosphatase (AP) and anti-AP which comprises the detection system of the amplified ELISA (a-ELISA) was investigated. The EIC appeared relatively homogeneous in sucrose density gradients and sedimented as a protein of 600-650 K daltons. Based on size and the results of double-label experiments, the EIC was shown to be composed of two moles of anti-AP and three moles of AP. During reaction with substrate at pH 9.6, greater than 50% of the AP is released as free enzyme and the released enzyme has the same activity as enzyme found in the EIC. The maximum yield of EIC is produced by solubilization of the antibody-AP equivalence precipitate with a 9-fold excess of the amount of AP required for precipitation at equivalence. EICs show no significant loss of activity when stored for one year at 4 degrees C, -20 degrees C or -70 degrees C. The EIC is most stable during long term storage (five years) in 50% glycerol at -20 degrees. Over the linear region of titration curve for dimeric and monomeric M315, the ratio of AP or EIC to M315 fails to show a constant stoichiometry. Using 131I-EIC and 125I-M315, it was determined that the lack of a constant stoichiometry in the linear region was due to differences in the amount of enzyme bound. Hence, stoichiometric quantitation of the primary antibody is not possible using the current a-ELISA.     

HLA-B alleles of the Navajo: no evidence for rapid evolution in the Nadene

New HLA-B locus alleles have been found in South American Amerindian populations but were largely absent in North American Amerindian tribes also descended from this first Paleo-Indian migration. We have now extended these studies to the Navajo, descendants of the second Nadene migration. No new functional alleles were found at the B locus of this tribe. This limited study supports the notion that while new B locus variants are common in South American Amerindians, it is more difficult to find new B locus alleles in North American native peoples. Whether this dichotomy is due to differences in pathogen environment and/or population structures between North and South America remains a subject of speculation.     

The ultrastructural features of aflatoxin B1-induced lesions in the rat liver

Hepatocellular carcinoma was induced in rats by administering aflatoxin B1 (AFB1) for 6 weeks. Malignant tumours were preceded by foci and nodules of altered hepatocytes. The ultrastructural characteristics of the nodular lesions have been studied and compared with those of the hepatocellular carcinoma cells. Alterations in the endoplasmic reticulum, junctional complexes and nuclei were common to both the basophilic and eosinophilic nodular cells and the carcinoma cells. These most likely represent hyperplastic changes rather than malignant alterations. The eosinophilic nodules were distinguished from other lesions by the abundance of concentric, membranous whorls in the cytoplasm of nodular cells. These cytoplasmic structures were also present in some hepatocellular carcinoma cells. The observations provided further evidence suggesting that the eosinophilic nodule, rather than the basophilic nodule, may play a role in the development of malignancy in the rat liver.     

Fluoride: an adjuvant for mucosal and systemic immunity.

Fluoride, the agent responsible for reduction of dental caries worldwide, and a recognized proliferative agent, is a potent adjuvant when given intragastrically to rats. Intragastric fluoride causes increases in the size and cellularity of the Peyer's patches and mesenteric lymph nodes as well as the number of plasma cells secreting IgG and IgA antibodies to ovalbumin given in their drinking water. Rats ingesting NaF and fed OA showed a significant increase in surface immunoglobulin expression on lymphocytes from the Peyer's patches and mesenteric lymph nodes. The frequency of CD4+ T cells in these lymphoid tissues was elevated while that of CD8+ T cells was significantly decreased. In separate experiments, rats parenterally immunized with myelin basic protein (MBP) and fed NaF twice weekly, had significantly elevated serum IgG antibody activity to MBP compared to similarly immunized rats not receiving NaF. The supplemental fluoride prescribed for infants and especially that which is inadvertently ingested by children and adults given fluoride gels, is within the concentration range of that which produced the effects we observed in rats. The adjuvant effect we describe thus has relevance for fluoride therapy worldwide.     

Antibiotic-resistant infections in primary care are symptomatic for longer and increase workload: outcomes for patients with E.coli UTIs

BACKGROUND: Antimicrobial resistance is considered to be one of the major threats to public health. However, the practical implications for patients and workload in primary care are largely unknown. AIM: To determine outcomes for patients managed in primary care with an antibiotic resistant compared to an antibiotic sensitive Escherichia coli (E. coli) urinary tract infection (UTI). DESIGN: Nested case control study with prospective measurement of outcomes. SETTING: Ten general practices in South Wales. METHOD: Patients consulting with symptoms suggestive of UTI identified through systematic sampling, and with a laboratory proven E. coli infection, were followed up by interview 1 month after their consultations and by searching of their medical records. RESULTS: Nine hundred and thirty-two patients were interviewed and had their medical records reviewed. The risk of patients reporting 'feeling poorly', 'frequency or pain on urinating' and being 'out of action' for more than 5 days after consulting was significantly increased for patients with resistant compared to sensitive infections. After adjusting for risk factors, there was an increased risk of 'frequency or pain on urinating' and 'being out of action' for those infected with a resistant E. coli. The median number of maximum reported days with at least one symptom was 12 days for patients with E. coli infections resistant to trimethoprim, 7 days for infections resistant to ampicillin, 7 days for infections resistant to any antibiotic, and 5 days for infections sensitive to all tested antibiotics. Even if treated with an appropriate antibiotic, infections caused by a resistant strain were symptomatic for longer. For those infected with an organism resistant to at least one antibiotic, the odds ratio (OR) for re-visiting their GP within the next 30 days for the UTI was 1.47 (95% confidence interval [CI] = 1.10 to 1.95). The OR was 1.49 (95% CI = 1.11 to 2.00) for ampicillin resistance and 2.48 (95% CI = 1.70 to 3.59) for trimethoprim resistance. CONCLUSIONS: Resistant E. coli UTIs are symptomatic for longer and cause increased work load in general practice.     

The effect of interplay interval on adjunctive behavior in humans in a game-playing situation

Thirty-seven undergraduate students played two consecutive games of backgammon where manipulated interplay intervals were either 10, 22.5, 45, or 90 sec in length. When visual access to the opponent's play was blocked by a curtain during the second game, rates of movement, drinking, and eating increased over baseline rates prevailing in the first game. Grooming rate was not affected. Functions relating behavior rate to interplay interval were not bitonic for any of the behaviors; however, response distributions for drinking demonstrated postplay peaks at each interplay interval. Problems of classifying human behaviors into adjunctive versus faculatative categories were discussed.     

Isotype commitment of human B cells that are transformed by Epstein-Barr virus.

Epstein-Barr virus (EBV) can transform a subpopulation of preactivated B cells thus promoting their growth and differentiation into plasma cells. In EBV-transformed clones of IgM-producing cells, the heavy chain constant region (CH) genes on the productive allele are fixed in germ-line configuration, whereas in isotype-switched clones the CH genes proximal to the expressed CH gene are deleted. In order to define more precisely the EBV-susceptible B cells, we sorted subpopulations of B cells on the basis of their cell surface Ig (sIg) isotypes, infected them with EBV, and determined which isotypes they could produce following transformation. Most precursors of IgM-producing plasma cells expressed both IgM and IgD on their surface, while a minority expressed IgM alone. Some B cell precursors of IgG- and IgA-producing cells also expressed sIgM, but surprisingly none expressed IgD. Those precursors of IgG and IgA producers, which bore sIgM, expressed it in relatively low levels, whereas B cells expressing high levels of sIgM were incapable of generating IgG and IgA producers. All of the precursors of IgG and IgA plasma cells expressed these isotypes on their cell surface. Interestingly, precursor B cells capable of producing the IgG3 and IgA2 subclasses could be respectively enriched on the basis of the presence or absence of cell sIgM. These results demonstrate the isotype precommitment of EBV-transformable B cells. They further suggest that residual IgM is transiently expressed on the surface of the IgG- and IgA-committed B cell precursors, whereas sIgD expression is extinguished earlier in the process of isotype switching via CH gene deletion.     

Interlaminar connections of tree shrew visual cortex.

The intracortical projections of neurons in layers II and upper III of tree shrew visual cortex were studied after terminal lesions in the supragranular layers of area 17. Examination for terminal degeneration was made using ultrastructural techniques. The majority of degenerating terminals were found in layers V and, to a lesser extent, VI, and were presynaptic to neural profiles in the following distribution: 80.5% on spines of small to medium size dendrites, 19% on dendrite shafts, and less than 1% on neuronal perikarya. Degenerating axons coursed in vertical bundles through layers III, IV, V and VI. These findings are similar to those previously described in rat visual cortex.