Monoclonal antibody detection of prolactin-binding subunits in the rabbit mammary gland.

The structure of prolactin (PRL) receptor in the rabbit mammary gland was examined using a receptor-specific monoclonal antibody (MAb). The PRL receptor preparation used was purified by making use of a PRL-affinity column. MAb inhibited the binding of PRL to the receptor, in a dose-dependent manner and completely at a high concentration. Using the receptor directly labelled by 125I, the preparation was incubated with MAbs and the immune complex was collected by Pansorbin and examined by SDS/polyacrylamide-gel electrophoresis. The autoradiography showed that three species with apparent Mr values of 77,000, 41,000 and 25,000 specifically reacted with MAbs. The pattern changed little in the presence or absence of dithiothreitol. Western blot analysis showed that two species (Mr 77,000 and 41,000) reacted with MAb. Affinity labelling of the receptor with labelled PRL revealed three bands with Mr values of 96,000, 60,000 and 43,000 on SDS gels. The high-Mr complex (Mr greater than 200,000) was always present at the top of the gel. These results show that the mammary gland contains at least three PRL-binding subunits. The differences in Mr before and after PRL binding were close to the Mr of PRL. This would suggest that each PRL binding subunit reacts with one PRL molecule.     

Merging Spline Approximations with Analysis of Nonlinear Regression Models

The paper outlines a method of analysis of intricically non-linear regression functions. The nonlinear model is first approximated by a cubic spline function. Thereafter, the standard methods of analysis in linear models are applied to obtain estimates and test statistics.     

Release of developmental constraints on tetrad shape is confirmed in inaperturate pollen of Potamogeton

Background and Aims

Microsporogenesis in monocots is often characterized by successive cytokinesis with centrifugal cell plate formation. Pollen grains in monocots are predominantly monosulcate, but variation occurs, including the lack of apertures. The aperture pattern can be determined by microsporogenesis features such as the tetrad shape and the last sites of callose deposition among the microspores. Potamogeton belongs to the early divergent Potamogetonaceae and possesses inaperturate pollen, a type of pollen for which it has been suggested that there is a release of the constraint on tetrad shape. This study aimed to investigate the microsporogenesis and the ultrastructure of pollen wall in species of Potamogeton in order to better understand the relationship between microsporogenesis features and the inaperturate condition.

Methods

The microsporogenesis was investigated using both light and epifluorescence microscopy. The ultrastructure of the pollen grain was studied using transmission electron microscopy.

Key Results

The cytokinesis is successive and formation of the intersporal callose wall is achieved by centrifugal cell plates, as a one-step process. The microspore tetrads were tetragonal, decussate, T-shaped and linear, except in P. pusillus, which showed less variation. This species also showed a callose ring in the microsporocyte, and some rhomboidal tetrads. In the mature pollen, the thickening observed in a broad area of the intine was here interpreted as an artefact.

Conclusions

The data support the view that there is a correlation between the inaperturate pollen production and the release of constraint on tetrad shape. However, in P. pusillus the tetrad shape may be constrained by a callose ring. It is also suggested that the lack of apertures in the pollen of Potamogeton may be due to the lack of specific sites on which callose deposition is completed. Moreover, inaperturate pollen of Potamogeton would be better classified as omniaperturate.Key words: Alismatales, callose, microsporogenesis, pollen aperture, Potamogeton illinoensis, P. polygonus, P. pusillus, tetrad shape     

Cellulase production on glucose-based media by the UV-irradiated mutants of Trichoderma reesei

From 22,791 mutants of a cellulase hyper-producing strain of Trichoderma reesei (Hypocrea jecorina), ATCC66589, as the parent, we selected two mutants, M2-1 and M3-1, that produce cellulases in media containing both cellulose and glucose. The mutation enabled the mutants to produce cellulases, which were measured as p-nitrophenyl β-d-lactopyranoside-hydrolyzing activities, in media with glucose as a sole carbon source, although M2-1 exhibited different sensitivities to glucose from M3-1. When the mutants were grown for 8 days on a medium with cellulose as a sole carbon source, the filter-paper-degrading activities (FPAs) per gram of cellulose were 257 and 281 U for M2-1 and M3-1, respectively, values that were 1.1–1.2 times higher than that of the parental strain. Cellulase production by M2-1 and M3-1 on a medium with a continuously fed mixture of glucose and cellobiose resulted in 214 and 210 U of FPA/gram carbon sources, respectively, whereas less efficient production (140 U of FPA/gram carbon source) was achieved by the parental strain. The improved cellulase productivity of the mutants allows us to use glucose as a carbon source for efficient on-site production of cellulases with quality/quantity-controlled feeding of soluble carbon sources and inducers.     

Surprising negative association between IgG1 allotype disparity and anti-adalimumab formation: a cohort study

Introduction

The human monoclonal antibody adalimumab is known to induce an anti-globulin response in some adalimumab-treated patients. Antibodies against adalimumab (AAA) are associated with non-response to treatment. Immunoglobulins, such as adalimumab, carry allotypes which represent slight differences in the amino acid sequences of the constant chains of an IgG molecule. Immunoglobulins with particular IgG (Gm) allotypes are racially distributed and could be immunogenic for individuals who do not express these allotypes. Therefore, we investigated whether a mismatch in IgG allotypes between adalimumab and IgG in adalimumab-treated patients is associated with the development of AAA.

Methods

This cohort study consisted of 250 adalimumab-treated rheumatoid arthritis (RA) patients. IgG allotypes were determined for adalimumab and for all patients. Anti-idiotype antibodies against adalimumab were measured with a regular radio immunoassay (RIA), and a newly developed bridging enzyme linked immunosorbent assay (ELISA) was used to measure anti-allotype antibodies against adalimumab. The association between AAA and the G1m3 and the G1m17 allotypes was determined. For differences between groups we used the independent or paired samples t-test, Mann-Whitney test or Chi square/Fisher's exact test as appropriate. To investigate the influence of confounders on the presence or absence of AAA a multiple logistic regression-analysis was used.

Results

Adalimumab carries the G1m17 allotype. No anti-allotype antibodies against adalimumab were detected. Thirty-nine out of 249 patients had anti-idiotype antibodies against adalimumab (16%). IgG allotypes of RA patients were associated with the frequency of AAA: patients homozygous for G1m17 had the highest frequency of AAA (41%), patients homozygous for G1m3 the lowest frequency (10%), and heterozygous patients' AAA frequency was 14% (P = 0.0001).

Conclusions

An allotype mismatch between adalimumab and IgG in adalimumab-treated patients did not lead to a higher frequency of AAA. On the contrary, patients who carried the same IgG allotype as present on the adalimumab IgG molecule, had the highest frequency of anti-adalimumab antibodies compared to patients whose IgG allotype differed from adalimumab. This suggests that the allotype of adalimumab may not be highly immunogenic. Furthermore, patients carrying the G1m17-allotype might be more prone to antibody responses.     

Exploiting the full power of temporal gene expression profiling through a new statistical test: Application to the analysis of muscular dystrophy data

Background  

The identification of biologically interesting genes in a temporal expression profiling dataset is challenging and complicated by high levels of experimental noise. Most statistical methods used in the literature do not fully exploit the temporal ordering in the dataset and are not suited to the case where temporal profiles are measured for a number of different biological conditions. We present a statistical test that makes explicit use of the temporal order in the data by fitting polynomial functions to the temporal profile of each gene and for each biological condition. A Hotelling T ²-statistic is derived to detect the genes for which the parameters of these polynomials are significantly different from each other.     

VanB-vanA incongruent VRE isolated from animals and humans in 1999

16 chicken isolates and four clinical isolates of VanB-vanA incongruent vancomycinresistant Enterococcus faecium strains without vanS were isolated in 1999. Pulsed-field gel electrophoresis revealed only a peripheral relationship between the chicken isolates and clinical isolates, but suggested clonal spread in the chicken isolates.     

Biodegradation of a polyvinyl alcohol-starch blend plastic film

Attempts were made to elucidate the degradation mechanism of a polyvinyl alcohol (PVA)-starch blend plastic. A part of the starch fraction of this plastic was dissolved into an aqueous phase in a control test. Treatment with a PVA-degrading bacterium or enzyme gave a maximal weight loss of approximately 70% and film breakage occurred. Since this plastic contains 40% PVA, it is apparent that not only the PVA fraction but also a considerable portion of the starch fraction was lost from the film by treatment with the PVA-degrading enzyme. As the PVA-degrading bacterium and enzyme used here showed no starch-degrading activity, loss of the starch fraction seems to depend on its dissolution with degradation of the PVA fraction. These experimental results indicated that the degradation of the PVA fraction is an important requisite for complete degradation or decomposition of this plastic film.     

Vancomycin-resistant enterococci in humans and imported chickens in Japan

The phenotypes and genotypes of 22 VanA-type vancomycin-resistant enterococci that had been isolated in Japan were examined. The VanA resistance determinant was plasmid mediated in each of the 22 strains. Of the 22 strains, 8 were isolated from different patients and 11 and 3 were obtained from different samples of chickens imported from Thailand and France, respectively. Three of the strains that were isolated from patients and the 11 strains isolated from the Thai chickens showed high-level vancomycin resistance (MICs, 512 to 1,024 micro g/ml) and low-level teicoplanin resistance (MICs, 0.5 to 4 micro g/ml). Each of these strains had three amino acid substitutions in the N-terminal region of the deduced VanS sequence. L50 was converted to V, E54 was converted to Q, and Q69 was converted to H compared to the vanS gene sequence of Tn1546.