A comparison of immunocytochemical staining enhancement methods using a rapid microtitre immunocytochemistry assay (MIA)

A method is described which allows rapid and quantitative comparison of immunocytochemical staining procedures. Cells grown and fixed in microtitre plates are probed with increasing dilutions of primary antibody and then stained using the procedures under test; the resulting staining intensities are determined using a microtitre plate reader. The microtitre immunocytochemistry assay (MIA) has been used to compare the sensitivities of enhancement procedures based on immunoperoxidase and immunogold staining. Silver enhancement of DAB staining was found to be the most sensitive technique giving up to 200 fold amplification of the peroxidase staining.     

Cryptococcal skin test antigen: preparation variables and characterization.

Antigen capable of eliciting delayed hypersensitivity reactions in the skin of sensitized guinea pigs could be extracted from Cryptococcus neoformans cells by stirring the cells from 3 to 5 days in concentrated urea or guanidine. Hydrolysis of urea to ammonia by cryptococcal urease accompanied urea extraction, but alkalinity appeared neither necessary nor sufficient for extraction. Antigen from live cells gave larger delayed skin reactions than did antigen from Formalin-killed cells. Peak skin test reactivity appeared to reside in protein-rich fraction having an elution volume on Sephadex G50 corresponding to a molecular weight of 10(4). Activity precipitated with half-saturated ammonium sulfate and could be detected in a single, narrow, rapidly migrating band on disc electrophoresis. Dialyzable proteinaceous antigen and high-molecular-weight, serologically active polysaccharide were present in the antigen, but not active in the delayed hypersensitivity reactions.     

Age related differences in binding of Concanavalin a to plasma membranes of isolated neurons

Neurons isolated from the lateral vestibular nucleus of young adult and senescent Fischer-344 rats were incubated with fluorescamine-labelled Concanavalin A (fl-Con A) alone, or following incubation in trypsin or Vibrio cholerae neuraminidase. They were then observed and photographed. Microdensitometric analysis of fluorescence micrographs showed that senescent rat neurons were significantly more fluorescent than those from young adult rats. Additionally, either patches or caps of fl-Con A were seen on the surface of neurons from senescent rats, while most young adult rat neurons bound fl-Con A uniformly. Pretreatment with trypsin or neuraminidase had no effect on the amount of fluorescence on the surface of senescent rat neurons, and only a slight effect on the surface distribution. Trypsin and neuraminidase treatments caused a significant increase in fluorescence on the neuronal plasma membranes of young adult rats and a rearrangement of the binding pattern in the majority of neurons observed.     

Emergence of influenza A H1N2 reassortant viruses in the human population during 2001

A recombinant vaccinia virus encoding rotavirus protein NSP3 driven by an internal ribosome entry site (IRES) from the encephalomyocarditis (EMC) virus was able to abate protein synthesis in BSC1 cells by 25-fold, with as much as 30% of the remaining protein synthesis being NSP3. Hence NSP3 shuts off host cell protein synthesis down to the level seen during rotavirus infection but is unable to prevent translation from EMC IRES-driven genes. This effect was abolished by deletions in the eIF4G-binding (aa 274-313) and the dimerization (aa 150-206) but not the viral mRNA-binding (aa 83-149) domains, supporting that NSP3 functions in vivo as a dimer. Binding of eIF4G by NSP3 has been implicated in interfering with mRNA 5'-3' circularization, hence such circularization is essential for translation in mammalian cells.     

Polymerase chain reaction for detection of herpesvirus simiae (B virus) in clinical specimens

Summary A polymerase chain reaction (PCR) was designed which is specific toMacaca fascicularis (cynomolgus monkey) isolates of B virus. The PCR primers produced the expected 188 basepair product from the Cyno 2 strain and seven other cynomolgus monkey isolates of B virus. Oligomer hybridization with a 31-mer oligonucleotide was used to confirm the origin of this product. The PCR failed to amplify DNA of Epstein-Barr virus, cytomegalovirus, varicella-zoster virus, and other alphaherpesviruses (herpes simplex virus types 1 and 2, four SA 8 isolates and three rhesus isolates of B virus). PCR testing of swabs obtained from four orally-infected cynomolgus monkeys confirmed the presence of B virus DNA in samples previously shown to be positive by culture. In addition, PCR detected B virus in several swabs from infected monkeys that were culture negative. Total DNA extracts from the trigeminal and sacral ganglia of these animals were tested by nested PCR and B virus DNA was detected in the trigeminal ganglia of 3 of the 4 orally-infected cynomolgus monkeys. Nested PCR did not detect B virus DNA in total DNA extracts obtained from the brains of the four monkeys.     

A model lightning safety policy for athletics

OBJECTIVE: The purpose of this paper is to present a model policy on lightning safety for athletic trainers. BACKGROUND: Among college athletic programs in the United States there is a serious lack of written policy on lightning safety. Available evidence shows that most National Collegiate Athletic Association (NCAA) Division I institutions, even though they are located in high lightning activity areas of the country, do not have formal, written lightning safety policies. CLINICAL ADVANTAGES/ RECOMMENDATIONS: The policy presented herein, which is at the forefront of such policies, is the lightning safety policy written as part of a policies and procedures manual for the division of sports medicine at a public NCAA Division I university. This is a policy based on practicality that utilizes the "flash-to- bang" method for determining the distance of lightning activity from the observer. The policy begins with the importance of prevention, including the daily monitoring of weather reports. The policy defines a "safe shelter" and specifies the chain of command for determining who removes a team or individuals from an athletic site in the event of dangerous lightning activity.     

Disruption of the ATM gene in breast cancer

Mutations in the ATM gene, which maps to 11q22-23, cause the multisystem recessive syndrome ataxia-telangiectasia (AT). Breast cancer has been reported in AT patients and carriers. Sporadic breast cancer is associated with loss of heterozygosity at or in the region of ATM and chromosomal abnormalities involving 11q23. We have investigated the chromosomes, nuclei and released chromatin fibers from nine primary breast carcinoma and eight cell lines by fluorescence in situ hybridization with four fluorochrome-labeled cosmids spanning the ATM gene. The ATM gene was disrupted in one primary breast carcinoma and in the cell lines MDA-MB-231 and MCF-7. The role of these aberrations in breast carcinomas, which may lead to gene dosage or dominant negative effects on gene function, requires further investigation.