Detection of Autographa californica and Heliothis zea baculovirus proteins by enzyme-linked immunosorbent assay (ELISA)

The structural proteins of Autographa californica (AcMNPV) and Heliothis zea (HzSNPV) nuclear polyhedrosis viruses were detected by indirect enzyme-linked immunosorbent assay (ELISA). The immunoassay detected less than 1 ng of AcMNPV protein. The extent of immunological relatedness between AcMNPV-occluded virus and AcMNPV polyhedral protein, AcMNPV-nonoccluded virus, Estigmene acrea granulosis virus, Amsacta moorei entomopoxvirus Heliothis zea NPV, and Lymantria dispar NPV was determined. No immunological relatedless was detected between HzSNPV, AcMNPV, and a persistent rod-shaped virus isolated from the Heliothis zea cell line (IMC-Hz-1). The polyhedral proteins of HzSNPV and AcMNPV were found to be immunologically identical.     

Colcemid treatment of myeloma prior to cell fusion increases the yield of hybridomas between myeloma and splenocyte

Effect of Colcemid treatment of myeloma (X63-Ag8-6.5.3.) prior to fusion with mouse spleen cell was studied in terms of hybridoma formation. Spleen cells from BALB/c mice immunized with various soluble antigens were fused with the myeloma cells by using polyethylene glycol solution. Colcemid treatment of myeloma cells prior to fusion increased the average number of hybridoma colonies per well by 26-570%. The yield of hybridomas producing antigen-specific antibodies was also higher with the Colcemid treatment. The results suggest that most of the proliferative hybridomas are formed by fusion of cells in the M-phase of the cell cycle.     

The solid phase attachment of fungal hyphae in an ELISA to screen for antifungal antibodies

A method to immobilize fungal hyphae onto the wells of 96-well microplates for use in an in-direct ELISA to screen for antifungal antibodies in sera and cell culture supernatants is described. The hyphae from three genera (Penicillium, Eurotium andFusarium) were successfully attached by overnight drying onto wells precoated with poly-L-lysine and glutaraldehyde. Microscopy revealed that the hyphae remained attached to the wells throughout the ELISA and antiserum titrations showed that the attached hyphae were uniformly coated and remained reactive. Background absorbances were low and the plates could be stored at –20 °C without loss of reactivity.     

Comparison of three ELISA kits for the differentiation of foot-and-mouth disease virus-infected from vaccinated animals

A study was performed to validate 3 FMDV 3ABC-I-ELISA kits developed in China for the differentiation of FMDV infected and vaccinated animals. Sets of sera from naive and vaccinated cattle as well as from cattle that had been infected were tested for antibodies against nonstructural proteins (NSPs) of FMDV by commercial diagnosis kits, Ceditest® FMDV-NS (Ceditest® kit), UBI® FMDV NONSTRUCTURAL PROTEIN ELISA DIRECTION INSERT (UBI® kit) and a FMDV 3ABC-I-ELISA kit developed at the Lanzhou Veterinary Research Institute. The test parameters (sensitivity and specificity) of the three kits were determined, and the result obtained from FMD 3ABC-I-ELISA kit was compared with that obtained from two foreign kits. The results indicated that the coincidence rate between the FMDV 3ABC-I-ELISA and Ceditest® kits was 98.05%, and the coincidence rate between the FMDV 3ABC-I-ELISA and UBI® kits was 94.4%; the sensitivity of both Ceditest® and FMDV 3ABC-I-ELISA kit was 100%. However, the sensitivity of the UBI® kit was only 81.8%. With sera from naive or vaccinated non-infected animals, the specificity of all tests exceeded 90%.     

Chemiluminescent immunoassay (CLIA) for the detection of brucellosis and tularaemia antigens

The detection of brucellosis and tularaemia infection agents is of particular interest for medical practice. The possibility of using enhanced chemiluminescence reactions for the determination of these agents is studied in this work. Light intensity depends on both the conjugate concentration used and the conditions at which the adsorption was performed. Optimal conditions for these test-systems were: ～ 20 μg/mL of Ig and 200 μg/mL (titre 1:20) of conjugate. As is seen from the chemiluminescent and spectrophotometric results the lowest determined concentrations are 10 and 30 ng/mL (for brucellosis) and 1 and 5 ng/mL (for tularaemia), respectively. Calibration curves in the antigen concentrations ranging from 10 to 2500 ng/mL (for brucellosis) and from 1 to 500 ng/mL (for tularaemia) are observed. Optical density depends linearly on the logarithm of the antigen concentration from 30 to 5000 ng/mL (for brucellosis) and from 5 to 250 ng/mL (for tularaemia). The results obtained permit the conclusion that the chemiluminescence method can be used in enzyme immunoanalysis for brucellosis and tularaemia antigens.     

Frequent overexpression of CADM1/IGSF4 in lung adenocarcinoma

We reported comprehensive screening for antigens (Ags) overexpressed on various carcinomas via isolation of human monoclonal antibodies (mAbs) that may be therapeutic in a previous paper (Proc. Natl. Acad. Sci. USA 105, 7287-7292, 2008). Twenty-one distinct Ags highly expressed on several carcinomas were identified and 356 mAbs with unique sequences turned out to bind to one of the 21 Ags. Among them CADM1/IGSF4 which had been originally referred to as tumor suppressor lung cancer 1 (TSLC1) was included. Therefore we examined the expression of CADM1 in lung cancers in this study. Eight different anti CADM1 mAbs were used for immunohistochemical analysis of 29 fresh lung cancer specimens. Staining patterns were categorized to six groups based on the extent of positive staining and the localization of stained portions. While overexpression of CADM1 was observed on the cell surface of adenocarcinomas at a high frequency, around 60%, positive stainings were rarely observed on that of other lung carcinomas including squamous cell carcinomas. Moreover, some clones among the eight mAbs gave different staining patterns from those by the other clones against the same fresh specimen, suggesting presence of variant forms of CADM1 differentiated by mAbs.     

Bt水稻田重要非靶标节肢动物权露于Cry2Aa蛋白的程度分析

根据风险=危险×机露的原理,在实验室条件下评价转基因作物对非靶标节肢动物影响时,所选择的代表性非靶标生物通常是在农田系统中较高地关露于转基因外源杀虫蛋白的节肢动物种.为了弄清Bt稻田主要节肢动物暴露于Cry蛋白的程度,选择合适的非靶标节肢动物,用于转基因抗虫水稻的风险评价,本文采用酶联免疫技术检测了水稻不同生长期从转cry2Aa基因水稻田采集的不同节肢动物体内Cry2Aa蛋白的含量.结果表明:不同节肢动物种体内的Cry蛋白含量差异显著.一些节肢动物体内不合Cry蛋白,而一些节肢动物体内含有较高的Cry蛋白;相对于花期后采集的节肢动物,在Bt水稻花期采集的节肢动物,特别是捕食性节肢动物体内的Cry蛋白含量较高;寄生性节肢动物体内未检测到Cry蛋白.这为在实验室条件下评价转基因水稻对农田非靶标节肢动物的影响奠定了基础.     

Prevalence, whole genome characterization and phylogenetic analysis of hepatitis B virus in captive orangutan and gibbon

Background Hepatitis B virus (HBV) is a public health problem worldwide and apart from infecting humans, HBV has been found in non‐human primates. Methods We subjected 93 non‐human primates comprising 12 species to ELISA screening for the serological markers HBsAg, antiHBs and antiHBc. Subsequently, we detected HBV DNA, sequenced the whole HBV genome and performed phylogenetic analysis. Results HBV infection was detected in gibbon (4/15) and orangutan (7/53). HBV DNA isolates from two gibbons and seven orangutans were chosen for complete genome amplification. We aligned the Pre‐S/S, Pre‐C/C and entire genomes with HBV sequences and performed phylogenetic analysis. The gibbon and orangutan viruses clustered within their respective groups. Conclusions Both geographic location and host species influence which HBV variants are found in gibbons and orangutans. Hence, HBV transmission between humans and non‐human primates might be a distinct possibility and additional studies will be required to further investigate this potential risk.     

Epidemiological Characteristics of Strongyloidiasis in Inhabitants of Indigenous Communities in Borneo Island,Malaysia

Epidemiological study on strongyloidiasis in humans is currently lacking in Malaysia. Thus, a cross-sectional study was carried out to determine the prevalence of Strongyloides stercoralis infection among the inhabitants of longhouse indigenous communities in Sarawak. A single stool and blood sample were collected from each participant and subjected to microscopy, serological and molecular techniques. Five species of intestinal parasites were identified by stool microscopy. None of the stool samples were positive for S. stercoralis. However, 11% of 236 serum samples were seropositive for strongyloidiasis. Further confirmation using molecular technique on stool samples of the seropositive individuals successfully amplified 5 samples, suggesting current active infections. The prevalence was significantly higher in adult males and tended to increase with age. S. stercoralis should no longer be neglected in any intestinal parasitic survey. Combination of more than 1 diagnostic technique is necessary to increase the likelihood of estimating the ‘true’ prevalence of S. stercoralis.