A simple and rapid microwave-assisted hematoxylin and eosin staining method using 1,1,1 trichloroethane as a dewaxing and a clearing agent

The use and practicability of microwave-assisted staining procedures in routine histopathology has been well established for more than 17 years. In the study reported here, we aimed to examine an alternative approach that would shorten the duration of dewaxing and clearing steps of hematoxylin and eosin (H & E) staining of paraffin sections by using a microwave oven. Although xylene is one of the most popular dewaxing and clearing agents, its flammability restricts its use in a microwave oven; thus we preferred 1,1,1 trichloroethane, which is not flammable, as the dewaxing and clearing agent in the present study. In Group I and Group II (control groups), intestine was processed with xylene and 1,1,1 trichloroethane, respectively. The sections were then stained with H & E according to the conventional staining protocol at room temperature and subdivided into two groups according to the duration of dewaxing and clearing in xylene. In Groups III and IV (experimental groups) similar tissues were processed with xylene and 1,1,1 trichloroethane, respectively; however, sections from these groups were divided into four subgroups to study the period required for dewaxing and clearing in 1,1,1 trichloroethane, then stained with H & E in the microwave oven at 360 W for 30 sec. Our conventional H & E staining procedure, which includes dewaxing, staining and clearing of sections, requires approximately 90 min, while our method using 1,1,1 trichloroethane and microwave heating required only 2 min. Our alternative method for H & E staining not only reduced the procedure time significantly, but also yielded staining quality equal or superior to those stained the conventional way. Our results suggest that 1,1,1 trichloroethane can be used effectively and safely as a dewaxing and clearing agent for H & E staining in a microwave oven.     

Characterization of 13 newly isolated strains of anaerobic, cellulolytic, thermophilic bacteria

Characteristics of 13 newly isolated thermophilic, anaerobic, and cellulolytic strains were compared with previously described strains of Clostridium thermocellum: ATCC 27405 and JW20 (ATCC 31549). Colony morphology, antibiotic sensitivity, fermentation end-products, and cellulose degradation were documented. All 13 strains were sensitive to erythromycin (5 μg/ml) and chloramphenicol (25 μg/ml), and all strains but one were sensitive to kanamycin (20 μg/ml). Polymerase chain reaction (PCR) amplification using primers based on gene sequences from C. thermocellum ATCC 27405 was successful for all 13 strains in the case of the hydrogenase gene and 11 strains in the case of phosphotransacetylase/acetate kinase genes. Ten strains amplified a product of the expected size with primers developed to be specific for C. thermocellum 16SrRNA primers. Two of the 13 strains did not amplify any product with the PCR primers designed for the phosphotransacetylase/acetate kinase and 16SrRNA primers. A MboI-like GATC- recognizing restriction activity was present in all of the five strains examined. The results of this study have several positive implications with respect to future development of a transformation system for cellulolytic thermophiles. Journal of Industrial Microbiology & Biotechnology (2001) 27, 275–280. Received 12 September 2000/ Accepted in revised form 20 November 2000     

A simple and rapid microwave-assisted hematoxylin and eosin staining method using 1,1,1 trichloroethane as a dewaxing and a clearing agent

The use and practicability of microwave-assisted staining procedures in routine histopathology has been well established for more than 17 years. In the study reported here, we aimed to examine an alternative approach that would shorten the duration of dewaxing and clearing steps of hematoxylin and eosin (H & E) staining of paraffin sections by using a microwave oven. Although xylene is one of the most popular dewaxing and clearing agents, its flammability restricts its use in a microwave oven; thus we preferred 1,1,1 trichloroethane, which is not flammable, as the dewaxing and clearing agent in the present study. In Group I and Group II (control groups), intestine was processed with xylene and 1,1,1 trichloroethane, respectively. The sections were then stained with H & E according to the conventional staining protocol at room temperature and subdivided into two groups according to the duration of dewaxing and clearing in xylene. In Groups III and IV (experimental groups) similar tissues were processed with xylene and 1,1,1 trichloroethane, respectively; however, sections from these groups were divided into four subgroups to study the period required for dewaxing and clearing in 1,1,1 trichloroethane, then stained with H & E in the microwave oven at 360 W for 30 sec. Our conventional H & E staining procedure, which includes dewaxing, staining and clearing of sections, requires approximately 90 min, while our method using 1,1,1 trichloroethane and microwave heating required only 2 min. Our alternative method for H & E staining not only reduced the procedure time significantly, but also yielded staining quality equal or superior to those stained the conventional way. Our results suggest that 1,1,1 trichloroethane can be used effectively and safely as a dewaxing and clearing agent for H & E staining in a microwave oven.     

Evolutionary rates for tuf genes in endosymbionts of aphids

The gene encoding elongation factor Tu (tuf) in aphid endosymbionts (genus Buchnera) evolves at rates of 1.3 x 10(-10) to 2.5 x 10(-10) nonsynonymous substitutions and 3.9 x 10(-9) to 8.0 x 10(-9) synonymous substitutions per position per year. These rates, which are at present among the most reliable substitution rates for protein-coding genes of bacteria, have been obtained by calibrating the nodes in the phylogenetic tree produced from the Buchnera EF-Tu sequences using divergence times for the corresponding ancestral aphid hosts. We also present data suggesting that the rates of nonsynonymous substitutions are significantly higher in the endosymbiont lineages than in the closely related free-living bacteria Escherichia coli and Salmonella typhimurium. Synonymous substitution rates for Buchnera approximate estimated mutation rates for E. coli and S. typhimurium, as expected if synonymous changes act as neutral mutations in Buchnera. We relate the observed differences in substitution frequencies to the absence of selective codon preferences in Buchnera and to the influence of Muller's ratchet on small asexual populations.      

PatternLab for proteomics: a tool for differential shotgun proteomics

Background  

A goal of proteomics is to distinguish between states of a biological system by identifying protein expression differences. Liu et al. demonstrated a method to perform semi-relative protein quantitation in shotgun proteomics data by correlating the number of tandem mass spectra obtained for each protein, or "spectral count", with its abundance in a mixture; however, two issues have remained open: how to normalize spectral counting data and how to efficiently pinpoint differences between profiles. Moreover, Chen et al. recently showed how to increase the number of identified proteins in shotgun proteomics by analyzing samples with different MS-compatible detergents while performing proteolytic digestion. The latter introduced new challenges as seen from the data analysis perspective, since replicate readings are not acquired.     

Characterization and potential evolutionary impact of transposable elements in the genome of Cochliobolus heterostrophus

Background

Cochliobolus heterostrophus is a dothideomycete that causes Southern Corn Leaf Blight disease. There are two races, race O and race T that differ by the absence (race O) and presence (race T) of ~ 1.2-Mb of DNA encoding genes responsible for the production of T-toxin, which makes race T much more virulent than race O. The presence of repetitive elements in fungal genomes is considered to be an important source of genetic variability between different species.

Results

A detailed analysis of class I and II TEs identified in the near complete genome sequence of race O was performed. In total in race O, 12 new families of transposons were identified. In silico evidence of recent activity was found for many of the transposons and analyses of expressed sequence tags (ESTs) demonstrated that these elements were actively transcribed. Various potentially active TEs were found near coding regions and may modify the expression and structure of these genes by acting as ectopic recombination sites. Transposons were found on scaffolds carrying polyketide synthase encoding genes, responsible for production of T-toxin in race T. Strong evidence of ectopic recombination was found, demonstrating that TEs can play an important role in the modulation of genome architecture of this species. The Repeat Induced Point mutation (RIP) silencing mechanism was shown to have high specificity in C. heterostrophus, acting only on transposons near coding regions.

Conclusions

New families of transposons were identified. In C. heterostrophus, the RIP silencing mechanism is efficient and selective. The co-localization of effector genes and TEs, therefore, exposes those genes to high rates of point mutations. This may accelerate the rate of evolution of these genes, providing a potential advantage for the host. Additionally, it was shown that ectopic recombination promoted by TEs appears to be the major event in the genome reorganization of this species and that a large number of elements are still potentially active. So, this study provides information about the potential impact of TEs on the evolution of C. heterostrophus.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-536) contains supplementary material, which is available to authorized users.     

Infection control and the significance of sputum and other respiratory secretions from adult patients with cystic fibrosis

Background

Although various hematologic abnormalities are seen in tuberculosis, immune thrombocytopenic purpura is a rare event.

Case Presentation

We report a case of a 29 year-old male who was presented with immune thrombocytopenia-induced hemoptysis, macroscopic hematuria and generalized petechiae. The patient was found to have clinical, microbiological and radiological evidence of active pulmonary tuberculosis. The immune thrombocytopenic purpura was successfully treated with anti-tuberculous drugs combined with corticosteroids and high dose immune globulin therapy.

Conclusion

Immune thrombocytopenic purpura can be one of the hematological manifestations of tuberculosis which has a global prevalence with increasing incidence secondary to HIV infection.     

Array2BIO: from microarray expression data to functional annotation of co-regulated genes

Background  

There are several isolated tools for partial analysis of microarray expression data. To provide an integrative, easy-to-use and automated toolkit for the analysis of Affymetrix microarray expression data we have developed Array2BIO, an application that couples several analytical methods into a single web based utility.