Orcein and Elastic Fibers

Orcein was first proposed as an elastic fiber stain by Taenzer in 1890, and has proved very useful for the purpose. It is an important constituent of the author's “Quad” stain. Unfortunately not all orcein samples have proved equally satisfactory, whether they are derived from the lichens from which the dye was originally prepared or have been manufactured by a synthetic process. At the present time several brands are available, and two of the brands of the synthetic product investigated have not proved to be the same thing; one of the latter proves only fair as an elastin stain, the other is one of the best samples the author has ever tried.     

Histochemical demonstration of collagen fibers in ascorbic-acid-fed cell cultures

Summary Nine cultures of fibroblast cell types and 13 epithelial-like cell types were maintained for 1 week in media supplemented with L-asborbic acid (50 μg per ml). All fibroblast-like cultures produced extracellular fibers that stained positively by a silver-impregnation reticulin stain. Nine of the 13 epithelial-like cultures produced fibers that stained positively for reticulin. Nearly all cultures not supplemented with ascorbic acid showed no fiber staining. Those few lines that stained positively for reticulin in the absence of ascorbic-acid supplementation demonstrated only slight reticulin formation. Reticulin from one fibroblast culture and one epithelial culture was examined by electron microscopy, and the silver-impregnated fibrils were morphologically identical to collagen. The reticulin was digestible with collagenase, providing further evidence that the silver-impregnation reticulin stain identifies collagen in culture. The demonstration of collagen can be performed easily in histology laboratories using Formalin-fixed cells, and provides a means of assaying a functional property of cells in culture which is characteristic of connective tissue fibroblasts in general as well as certain specialized epithelia.     

Complete Staining of Nerve Fiber and Myoneural Junctions with Acetylthiocholine and Silver

We describe a combined stain for simultaneous demonstration of the preterminal axons and cholinesterase activity at myoneural junctions of mammalian muscles. This technique employs acetylthiocholine iodide as the substrate for cholinesterase activity and silver nitrate impregnation of preterminal axons. The procedure is rapid, simple and Uses fresh muscles. Intramuscular nerves, preterminal axons and myoneural junctions are stained simultaneously brown or black with minimal background staining of connective tissue and muscle fibers.     

Modification of the Silver Proteinate Impregnation Technique for Protozoa and Cultured Nerve Cells

Silver impregnation with silver-protein compounds is widely used for staining tissue sections and cell cultures. Some authors report that the results obtained with these methods have not always been reproducible because the reagent's composition varies according to the manufacturer. To avoid this problem in the method described in this paper, a silver proteinate, produced in our own laboratory is used. Although our method is based on Bodian's, the modifications we have made allows its use for both free-living cells (protozoa) and cells grown in culture (nerve cells). The significant modifications are 1) different fixation, 2) postfixation with Cajal's for-mol-bromide, 3) changes in the duration of the impregnation steps technique and 4) elimination of metallic copper. The method reported here enables us to use silver proteinate whenever we require it and to control the composition of the silver proteinate. This technique can be used for cells cultured in either plastic or glass.     

Complete Staining of Nerve Fiber and Myoneural Junctions with Acetylthiocholine and Silver

We describe a combined stain for simultaneous demonstration of the preterminal axons and cholinesterase activity at myoneural junctions of mammalian muscles. This technique employs acetylthiocholine iodide as the substrate for cholinesterase activity and silver nitrate impregnation of preterminal axons. The procedure is rapid, simple and Uses fresh muscles. Intramuscular nerves, preterminal axons and myoneural junctions are stained simultaneously brown or black with minimal background staining of connective tissue and muscle fibers.     

Molecular traceability of beef from synthetic Mexican bovine breeds

Traceability ensures a link between carcass, quarters or cuts of beef and the individual animal or the group of animals from which they are derived. Meat traceability is an essential tool for successful identification and recall of contaminated products from the market during a food crisis. Meat traceability is also extremely important for protection and value enhancement of good-quality brands. Molecular meat traceability would allow verification of conventional methods used for beef tracing in synthetic Mexican bovine breeds. We evaluated a set of 11 microsatellites for their ability to identify animals belonging to these synthetic breeds, Brangus and Charolais/Brahman (78 animals). Seven microsatellite markers allowed sample discrimination with a match probability, defined as the probability of finding two individuals sharing by chance the same genotypic profile, of 10(-8). The practical application of the marker set was evaluated by testing eight samples from carcasses and pieces of meat at the slaughterhouse and at the point of sale. The DNA profiles of the two samples obtained at these two different points in the production-commercialization chain always proved that they came from the same animal.     

A Comparison of American Stains with Recent German Products

In 1947, 31 samples of stains of German manufacture were collected in that country, with the cooperation of the U. S. Department of Commerce and one of the American stain manufacturers. They have been compared with current American products by the same tests as used for the samples submitted for certification. Only two proved distinctly superior to the corresponding American dyes, three others as good as the best American samples, four proved entirely unsatisfactory, while seven others were abnormal from the spectrophotometric standpoint although performing satisfactorily when used in the staining procedures by which they were tested.     

Microbiological quality of bottled water sold in retail outlets in Nigeria.

The microbiological quality of four brands of bottled water sold in retail outlets in Nigeria were assessed by routine methods in 90 samples. Samples of two brands were acidic in the pH range 3.5-5.9. Faecal coliforms and streptococci were not recovered from any sample. Heterotrophic plate counts (HPC) numbered 50-800 cfu/ml in two brands, A and B, and 100-87,000 cfu/ml in C and D. Component colony types among the HPC bacteria in brands C and D produced water-soluble, fluorescent pigments on colony count and other agar media, and occurred in 11 of 16 batches: their numbers varied from 60 to 82,000 cfu/ml. Presumptive antibiotic-resistant proportions of the HPC bacteria were also recovered from brands C and D on agar amended with ampicillin, penicillin or streptomycin. Five isolates from the green pigmented colonies, representing five batches of brands C and D were satisfactorily identified as strains of Pseudomonas aeruginosa. These strains resisted between four and nine of 14 antibiotics tested. Resistance to tetracycline was eliminated in one isolate when it was cured by incubation at 42 degrees C for 100 h, but gel electrophoretic resolution of the DNA did not reveal presence of a plasmid. Strains of Bacillus were the second most commonly isolated bacteria; they were the only colony types in most samples with low HPC counts.     

Differentiation of Synthetic from Biogenic Raw Materials in Various Foods with a Liquid Scintillation Counter

By estimating the ¹⁴C content of many acetic acid samples prepared from vinegars, Worcester sauces, ketchups and pickles with a liquid scintillation counter, it was proved that synthetic acetic acid mixed with biogenic one could be discriminated with considerable accuracy. It was also found that several products obtained from the open market contained synthetic acetic acid though they were represented to have been prepared exclusively from fermentation vinegar.     

Microbiological quality of bottled water sold in retail outlets in Nigeria

M.T. OGAN. 1992. The microbiological quality of four brands of bottled water sold in retail outlets in Nigeria were assessed by routine methods in 90 samples. Samples of two brands were acidic in the pH range 3.5–5.9. Faecal coliforms and streptococci were not recovered from any sample. Heterotrophic plate counts (HPC) numbered 50–800 cfu/ml in two brands, A and B, and 100–87000 cfu/ml in C and D. Component colony types among the HPC bacteria in brands C and D produced water-soluble, fluorescent pigments on colony count and other agar media, and occurred in 11 of 16 batches: their numbers varied from 60 to 82000 cfu/ml. Presumptive antibiotic-resistant proportions of the HPC bacteria were also recovered from brands C and D on agar amended with ampicillin, penicillin or streptomycin. Five isolates from the green pigmented colonies, representing five batches of brands C and D were satisfactorily identified as strains of Pseudomonas aeruginosa. These strains resisted between four and nine of 14 antibiotics tested. Resistance to tetracycline was eliminated in one isolate when it was cured by incubation at 42C for 100 h, but gel electrophoretic resolution of the DNA did not reveal presence of a plasmid. Strains of Bacillus were the second most commonly isolated bacteria; they were the only colony types in most samples with low HPC counts.     

A Modified Silver Impregnation Technique for the Observation of Thick Sections of Lymph Nodes Perfused with Colloidal Carbon

For many years, a variant of the silver impregnation technique of Bielchowsky has been used to study the lymph node because it clearly outlines the various structures which are usually hard to contrast with standard staining methods. Like other variants of silver impregnation, this method blackens the cell nuclei as well as the reticular fibers; however, it inhibits the impregnation of the nuclear chromatin immediately adjacent to fibers. Hence, this variant selectively darkens the lymphoid cell populations of the nodal structures which contain a loose fiber network.

To study the blood vascular network of the lymph node based on perfusion of colloidal carbon, a staining procedure was needed which would contrast nodal structures on thick sections, while allowing the carbon-filled small blood vessels to be distinguished from the impregnated coarse reticular fibers. In an attempt to adapt this variant of Bielchowsky's technique, 10, 20, 40 and 60 nm thick sections from rat nodes, fixed in a solution of Bouin-Hollande for 72 hr, were silver impregnated with serial dilutions (1:2 to 1:128) of the ammoniacal silver solution. Forty-micrometer thick sections impregnated with a 1:16 dilution of the original silver solution at 37 C and for 30 min provided the best results for the conditions.     

微量蛋白检测中不同银染方法的比较与分析

随着生物化学技术的不断发展,作为检测SDS-聚丙烯酰胺凝胶电泳(SDS-PAGE)中微量蛋白的银染方法也在不断改进和发展.采用4种不同的银染方法检测不同含量的牛血清白蛋白,结果显示单纯的银染过程中如果使用戊二醛固定会使蛋白检出更快速灵敏,而结合考马斯亮蓝的复合银染则较单纯银染灵敏度提高了5～7个数量级.     

Microwaves Applied to Silver Impregnations with Ammoniacal Silver Carbonate

Techniques for impregnation with ammoniacal silver carbonate provide valuable information on all types of tissue; however, the time investment required to impregnate a few sections has limited their application. We have shortened the impregnation times by using microwaves in techniques for reticular fibers, astrocytes, nerve fibers and chromaffin cells. The results were satisfactory with markedly reduced impregnation time and elimination of nonspecific silver deposits.     

Infraciliature and morphogenesis in three rumen Diplodinium ciliates, Diplodinium polygonale, Diplodinium leche, and Diplodinium nanum, observed by light microscopy

Infraciliature and morphogenesis of three rumen ophryoscolecid ciliates, Diplodinium polygonale Dogiel, 1925, Diplodinium leche Imai et al., 1992, and Diplodinium nanum Imai, 1988, are described from pyridinated silver carbonate-impregnated specimens. These three species have two polybrachykineties in the buccal area and a polybrachykinety in the dorsal ciliary zone. The vestibular polybrachykinety (VP) of D. polygonale and D. leche arises from the dorsal extremity of the adoral polybrachykinety (AP) as in Entodinium species, extending toward the left in D. polygonale and toward the left posterior in D. leche. The VP of D. nanum arises from the inner side of the AP, separate from its dorsal extremity, as in other Diplodinium species and extends toward the left posterior. These series of the polybrachykinety arrangements in D. polygonale, D. leche, and D. nanum can be regarded as transitional forms in the evolution of an Entodinium-like ancestor to Diplodinium. Morphogenesis of these three Diplodinium species is not different from that of other Diplodinium species.     

蛋白质双向电泳双胺染色方法的改进

讨论了蛋白质双向电泳检测的各种方法,并对本实验室使用过的两种银染方法（非双胺银染及双胺银染）进行对比研究。发现本实验室改进的双胺银染色方法具有以下特点：（1）背景异常清晰,蛋白质与底色反差大;（2）敏感度大大提高,检测最低蛋白质量可达fg级,对微量表达的蛋白质具有极好分离效果。能清晰地检测出低丰度表达的蛋白质。     

A Simplified Nissl Stain with Thionin

Recently two articles on the use of thionin as a cell stain for neurological materials have appeared. One utilizes a solution buffered in the acid range³; the other uses a “steaming” staining solution⁴. For some time we have been using thionin as a routine stain after either formalin or alcohol fixation and our method is so simple and has given such satisfactory results with a variety of brands of thionin that it seemed to be worthy of more general use. Briefly the method consists of placing the celloidin sections in a 0.05% solution of Li₂CO₃ (the percentage of Li₂CO₃ is non-critical) for about 5 minutes and then grossly overstaining in a 0.25% solution of thionin in a 0.05% solution of Li₂CO₃ in distilled water. The overstaining is necessary if all the stain is to be removed from the background. The sections are then passed through distilled water, 70 or 80% alcohol, two changes of butyl alcohol, two changes of xylene and mounted with Clarite. For most material, split mica cover-slips are quite satisfactory. The time of differentiation may be considerably lessened by the use of the differentiator recommended by Neumann (1942) except that we find the chloroform superfluous and transfer the sections to the aniline solution from 95% alcohol. Less fading seems to occur if the aniline differentiator is followed by a saturated solution of Li₂CO₃ in 95% alcohol.     

Staining Protozoa with Janus Green B

The use of janus green to stain mitochondria has long been known. While using it to study the mitochondria in Trichomonas buccalis it was found to stain the flagella also. This is an easy and quick method of determining the number of anterior flagella of the trichomonads and so of distinguishing the trichomonads from the pentatrichomonads in intestinal infections. This stain proved so useful as a flagella stain that it was applied to numerous protozoa with interesting results.     

Contamination of microbial foreign bodies in bottled mineral water in Tokyo,Japan

A total of 292 imported and domestic bottled mineral waters (90 brands) obtained from consumers and retailers were examined, by eye, for observable microbial foreign bodies. Fungal and bacterial foreign contaminants were found in 45 samples of water (20 brands) and in 14 samples of water (10 brands), respectively. Of the samples of water found to be contaminated, 41 (22 brands) were imported and 18 (8 brands) were produced domestically. Of 22 brands that were contaminated, 20 (91%) had been sterilized by at least one method. Forty-eight (98%) of 49 samples confirmed with foreign bodies were less than 1 year old. Among the moulds isolated the most predominant genus was Penicillium, followed by Acremonium and Cladosporium. The samples that contained fungi were less contaminated by bacteria than those that contained observable bacterial foreign bodies.     

LIBS: A Quality Control Tool for Food Supplements

In the present paper the ability of calibration free laser induced breakdown spectroscopy (CF-LIBS) as a quality control tool to monitor the composition of different minerals present in food supplement samples belonging to Indian brands (brand-A and brand-B) has been demonstrated. LIBS spectra of these two food supplements (brand-A and brand-B) available in the form of tablet have been recorded. As reported by manufacturers of these two food supplements, LIBS spectra of brand-A contains the spectral signatures of minerals like Ca, Mg, C, P, Zn, Fe, Cu, and Cr whereas LIBS spectra of brand-B shows the presence of spectral lines like Ca, Mg and C. The spectral signatures of Na and K are also found in both brands whereas spectral signature of Ti is observed only in brand-B but these elements are not mentioned on the nutritional label of the brands. The quantitative analysis of mineral contents in food supplements has been done using CF-LIBS for brand A and brand B to verify the content of the minerals reported by the manufacturer of the food supplements. Our results show that Ca and Mg are the main matrix elements of these brands. The concentration of minor and trace elements estimated using CF-LIBS technique is found in agreement with the reported nutritional values of both the brands. The concentration of major elements Ca and Mg are also estimated from Atomic Absorption Spectroscopy which is in close agreement with CF-LIBS result.     

Improved immunological membrane filter method for detection of food-borne Salmonella strains.

An improved membrane filter method that involves the use of an enzyme-labeled antibody stain has been developed for the rapid detection of Salmonella species in foods. The procedure is carried out directly on a hydrophobic grid-membrane filter without requiring transfer by blotting to nitrocellulose. Pure cultures of 54 Salmonella species and 10 foods artificially contaminated with Salmonella colindale gave a positive reaction in which Salmonella colonies were visible as purple dots. Of 11 nonsalmonella organisms, only Citrobacter freundii reacted with Spicer-Edwards antiserum. Of 22 naturally contaminated food samples, 10 were positive for both the hydrophobic grid-membrane filter procedure of the Association of Official Analytical Chemists and the improved enzyme-labeled antibody stain method, and there was perfect agreement between the methods. Of these 10 positive samples, one was negative by the Health Protection Branch method; of the negative samples, two were positive by this latter method. The improved enzyme-labeled antibody stain method allows detection of Salmonella spp. in foods within 48 h, requires little equipment, and is inexpensive, easy to perform, and suitable for automated detection.