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PubMed2000

Background-free, high sensitivity staining of proteins in one- and two-dimensional sodium dodecyl sulfate-polyacrylamide gels using a luminescent ruthenium complex.

SYPRO Ruby dye is a permanent stain comprised of ruthenium as part of an organic complex that interacts noncovalently with proteins. SYPRO Ruby Protein Gel Stain provides a sensitive, gentle, fluorescence-based method for detecting proteins in one-dimensional and two-dimensional sodium dodecyl sulfate-polyacrylamide gels. Proteins are fixed, stained from 3h to overnight and then rinsed in deionized water or dilute methanol/acetic acid solution for 30 min. The stain can be visualized using a wide range of excitation sources commonly used in image analysis systems including a 302 nm UV-B transilluminator, 473 nm second harmonic generation (SHG) laser, 488 nm argon-ion laser, 532 nm yttrium-aluminum-garnet (YAG) laser, xenon arc lamp, blue fluorescent light bulb or blue light-emitting diode (LED). The sensitivity of SYPRO Ruby Protein Gel Stain is superior to colloidal Coomassie Brilliant Blue (CBB) stain or monobromobimane labeling and comparable with the highest sensitivity silver or zinc-imidazole staining procedures available. The linear dynamic range of SYPRO Ruby Protein Gel stain extends over three orders of magnitude, which is vastly superior to silver, zinc-imidazole, monobromobimane and CBB stain. The fluorescent stain does not contain superfluous chemicals (formaldehyde, glutaraldehyde, Tween-20) that frequently interfere with peptide identification in mass spectrometry. While peptide mass profiles are severely altered in protein samples prelabeled with monobromobimane, successful identification of proteins by peptide mass profiling using matrix-assisted laser desorption/ionization mass spectrometry was easily performed after protein detection with SYPRO Ruby Protein Gel stain.

Dextrans↗

PubMed1984

The distribution of Timm's stain in the nonsulphide-perfused human hippocampal formation.

A detailed description is given of the distribution of Timm's staining in the human hippocampal formation. In sections cut transverse to the long axis of the hippocampal formation the same pattern of staining is seen throughout almost all of the structure, i.e., through all the main body except the most caudal region and, following the medial bend made by the long axis, through most of the pes hippocampi. The staining typically filled almost the whole of the hilus of the dentate gyrus and the whole depth of the adjacent stratum pyramidale and stratum lucidum of subfield CA3. Rostrally, staining was traced into the tip of the uncus. Caudally, staining was traced to the inferior surface of the splenium at the apex of the caudal taper of the hippocampus. No staining was found in the gyrus fasciolaris, indusium griseum, or anterior hippocampal rudiment. Except in the medial part of the pes hippocampi, the Timm's staining did not reach the proximal border of the subfield CA1. In some sections of the main body of the hippocampus a narrow, infragranular stain-free zone was observed in the hilus. In the dentate gyrus of three older subjects, but not in those of three younger subjects, supragranular staining was found. It is argued that the Timm's staining described in this study is specific to the hippocampal mossy fibres. The distribution of the staining is discussed in relation to the boundaries of subfields CA3 and CA2 as delineated by other authors.

Aged↗

PubMed1994

Biotin staining in the giant fiber systems of the lobster.

The avidin-biotin-complex method is a popular immunocytochemical technique. This method labels consistently a group of neurons in the lobster ventral nerve cord in the absence of primary antibodies. The specific staining is due to a relatively high level of endogenous biotin (or biocytin) in these neurons. These biotin-positive neurons are located in the supraesophageal, thoracic, and abdominal ganglia. Intraaxonal injection of Lucifer yellow followed by Texas red-conjugated streptavidin staining reveals that the neurons are members of the medial giant (MG) and lateral giant (LG) systems, which are important in mediating rapid tail flipping during escape maneuvers. In neuronal somata, staining is restricted to the cytoplasm. Within MG axons, staining appears as punctate, subaxolemmal structures. Preincubating nerve cords in biocytin or direct intraaxonal injection of biocytin enhances staining of these punctate organelles. In LG axons, staining is localized to fragments of braided filamentous structures that also appear to be associated with the axolemma. Preincubation of ventral nerve cords in various concentrations of biocytin results in the appearance of additional groups of stained neurons, suggesting that there are subsets of neurons with specific biocytin-uptake or -retention mechanisms. In the crayfish, biotin-positive staining is confined to the MG neurons; the LG neurons are not stained. In the earthworm, no staining is observed in the MG and LG axon escape systems. In the goldfish, no biotin-staining is seen in the Mauthner neurons and their axons. The significance of specific localization of biotin or biocytin to subsets of neurons is unclear. It may reflect the presence of high levels of biocytin moieties on biotin-dependent enzymes. Biotin is an important cofactor in the catalytic functions of several decarboxylases crucial in energy production and lipogenesis. Axons of the giant fiber systems in lobsters and crayfish may have high energy and fatty acid synthesis requirements. Increased levels of biotin accumulation may also be related to other functions of the giant axon systems, such as the formation of electrical synapses among themselves and with phasic motoneurons.

Animals↗

PubMed1976

Nuclear stains with soluble metachrome metal mordant dye lakes. The effect of chemical endgroup blocking reactions and the artificial introduction of acid groups into tissues.

Following our study on the effect of deoxyribonucleic acid (DNA) extraction on nuclear staining with soluble metal mordant dye lakes covering 29 dye lakes we chose a series of lakes representing the three groups: (1) readily prevented by DNA removal, (2) weakened by DNA extraction but not prevented, (3) unaffected by DNA removal, for application of other endgroup blockade reactions. The lakes selected were alum and iron hematoxylins, iron alum and ferrous sulfate galleins, Fe2+ gallo blue E, iron alum celestin blue B, iron alum fluorone black and the phenocyanin TC-FeSO4 sequence. Azure A with and without an eosin B neutral stain, was used as a simple cationic (and anionic) dye control. Methylation was less effective than with simple cationic dyes, but did weaken celestin blue, gallo blue E and phenocyanin Fe2+ nuclear stains. These dyes also demonstrate other acid groups: acid mucins, cartilage matrix, mast cells, central nervous corpora amylacea and artificially introduced carboxyl, sulfuric and sulfonic acid groups. Alum hematoxylin stained cartilage weakly and demonstrated sulfation and sulfonation sites. The iron galleins, iron fluorone black and acid iron hematoxylin do not. A pH 4 iron alum hematoxylin gave no staining of these sites; an alum hematoxylin acidified with 1% 12 N HCl gave weaker results. Deamination prevented eosin and orange G counterstains but did not impair nuclear stains with any of the mordant dye lakes. The simple acetylations likewise did not alter mordant dye nuclear staining, the Skraup reagent gave its usual sulfation effect on other tissue elements, but did not alter nuclear stains by mordant dyes. The mordant dyes do not bind to periodic acid engendered aldehyde sites and p-toluidine/acetic acid and borohydride aldehyde blockades did not alter mordant dye lake nuclear staining. Nitration by tetranitromethane, which blocks azo coupling of tyrosine residues, did not alter nuclear staining by the mordant dye lakes. Benzil at pH 13, which prevents the beta-naphthoquinone-4-Na sulfonate (NQS) arginine reaction and the Fullmer reaction of basic nucleoprotein, did not affect iron gallein, iron or alum hematoxylin stains of nuclei or lingual keratohyalin.

Aldehydes↗

PubMed1983

An investigation of aldehyde fuchsin staining of unoxidized insulin.

Aldehyde fuchsin is a standard stain for the secretion granules of pancreatic B cells. The participation of either insulin or proinsulin in aldehyde fuchsin staining is in dispute. There is some evidence that permanganate oxidized insulin is stained by aldehyde fuchsin. Aldehyde fuchsin staining of unoxidized insulin has not been investigated adequately despite excellent staining results with tissue sections. Unoxidized insulin and proinsulin suspended by electrophoresis in polyacrylamide gels were fixed with Bouin's fluid and placed in aldehyde fuchsin for one hour. Because the unoxidized proteins were not stained by aldehyde fuchsin, it was concluded that neither insulin or proinsulin are responsible for the intense aldehyde fuchsin staining of unoxidized pancreatic B cell granules in tissue sections. A series of controlled experiments was undertaken to test the effects of fixatives, oxidation and destaining procedures on aldehyde fuchsin staining of insulin, proinsulin and other proteins immobilized in polyacrylamide gels. It was demonstrated that only oxidized proteins were stained by aldehyde fuchsin and that cystine content of the proteins had no apparent relation to aldehyde fuchsin staining. It was concluded that neither insulin nor proinsulin is likely to be responsible for the intense aldehyde fuchsin staining of unoxidized pancreatic B cell granules in tissue sections.

Animals↗

PubMed1980

Histological staining of pre- and postsynaptic components of amphibian neuromuscular junctions.

We have developed a combined staining technique whereby pre- and postsynaptic structures of the neuromuscular junction can be simultaneously visualized in the light microscope. The general approach was first to stain presynaptic nerve terminals using nitroblue tetrazolium (NBT), which when reduced to its diformazan state coloured the entire nerve terminal arborization blue. When the NBT stain was combined with the Karnovsky acetylcholinesterase (AChE) procedure the blue-coloured nerve terminal processes were vividly outlined by the brown copper ferrocyanide-AChE reaction product. Experiments were performed to ensure that the NBT AChE method stained nerve reliably and that even extremely small neural processes were stained and visualized in the light microscope. This method stained cholinergic neuromuscular junctions and unmyelinated axons in a variety of preparations. Aldehyde pre-and poststaining fixation markedly affected the quality of NBT nerve stain. In addition, glutaraldehyde had a direct role in the staining process. The quality of the nerve terminal staining was affected by the pH and the constituents of the staining solution. The powerful experimental advantage of the NBT-AChE stain for neuromuscular junctions resulted from the sharp colour contrast which made possible accurate determinations of the relationship between the presynaptic nerve terminal arborization and the postsynaptic junctional AChE activity.

Acetylcholinesterase↗

PubMed1976

The effect of graded 60 degrees C 1N nitric acid extraction and of deoxyribonuclease digestion on nuclear staining by metachrome mordant dye metal salt mixtures.

We can divide metachrome mordant staining of nuclei after graded 60 degrees C 1 N nitric acid extraction into three groups. The Feulgen nucleal reaction and dilute cationic dye staining of nuclei are abolished in about 30 minutes. With one group of metachrome dyes nuclear staining is lost with acid exposures of one hour or less. In a second group nuclear staining is weakened by 30-60 minute extractions, but persists in recognizable grade for 4-6 hours. In the third group nuclear staining remains almost unimpaired for 4-6 hours. In the first group the nuclear staining seems clearly assignable to the nucleic acids and to DNA in particular. In the second group loss of part of the reactivity on short exposure indicates some participation of DNA in the control staining result, as well as participation of basic nucleoprotein. In the third group staining seems assignable largely to basic nucleoprotein. The five gallocyanin group dyes, all in group 1, all possess a dialkylamino group, probably functioning as an ammonium chloride.Hematoxylin, the flurone blacks and gallein all present an o-hydroxysemiquinone group which probably acts as a weak acid, in addition to the carboxyl group of gallein which gives the strongest staining of nuclei at the longest acid exposure. Deoxyribonuclease digestion (2 hours, 37 degrees C) separated sharply a class in which nuclear staining failed completely, a class in which nuclear staining was fully equal to that in the control preparations and an intermediate group in which slight, moderate, or severa impairment was present. Generally there was good agreement between the two methods of nucleic acid removal, despite the fixation difference. In each case, however, the extraction procedure was one worked out for the fixation on which it was used.

Alum Compounds↗

PubMed1999

Gram staining with an automatic machine.

This study was undertaken to develop a new Gram-staining machine controlled by a micro-controller and to investigate the quality of slides that were stained in the machine. The machine was designed and produced by the authors. It uses standard 220 V AC. Staining, washing, and drying periods are controlled by a timer built in the micro-controller. A software was made that contains a certain algorithm and time intervals for the staining mode. One-hundred and forty smears were prepared from Escherichia coli, Staphylococcus aureus, Neisseria sp., blood culture, trypticase soy broth, direct pus and sputum smears for comparison studies. Half of the slides in each group were stained with the machine, the other half by hand and then examined by four different microbiologists. Machine-stained slides had a higher clarity and less debris than the hand-stained slides (p < 0.05). In hand-stained slides, some Gram-positive organisms showed poor Gram-positive staining features (p < 0.05). In conclusion, we suggest that Gram staining with the automatic machine increases the staining quality and helps to decrease the work load in a busy diagnostic laboratory.

Algorithms↗

PubMed1986

Effects of various agents on staining of the maturation pattern at the surface of rat incisor enamel.

In the maturation zone, two types of ameloblasts are arranged as bands across the rat incisor; these corresponded with a staining pattern at the surface which reflected the alternating pattern of ruffle-ended and smooth-ended ameloblasts. PAS stain showed bands and stripes similar to those following glyoxal bis(2-hydroxyanil) (GBHA) staining; these stains visualize the organic (PAS) and inorganic (GBHA) components of the maturation pattern. To further elucidate the nature of these bands, dissected rat incisors were treated with various agents prior to staining with GBHA or PAS. Guanidine for 2 h at room temperature showed no maturation pattern when stained with GBHA, as did teeth treated with EDTA and a bisophosphonate (HEBP). Hydrochloric acid and nitric acid removed the layer of outer enamel and incisors did not stain with GBHA, suggesting that the staining is a surface-related phenomenon. As staining was abolished by either the removal of mineral (EDTA) or protein (guanidine), the concurrent localization of non-crystal-bound calcium by GBHA staining, and of glycoprotein by PAS staining, indicates that calcium is associated with glycoprotein at the surface of the enamel.

Ameloblasts↗

PubMed1996

Nucleic acid stains as indicators of Giardia muris viability following cyst inactivation.

A reliable viability assay for Giardia is required for the development of disinfection process design criteria and pathogen monitoring by water treatment utilities. Surveys of single-staining nucleic acid dyes (stain dead parasites only), and double-staining vital dye kits from Molecular Probes (stain live and dead parasites) were conducted to assess the viability of untreated, heat-killed, and chemically inactivated Giardia muris cysts. Nucleic acid staining results were compared to those of in vitro excystation and animal infectivity. Nucleic acid stain, designated as SYTO-9, was considered the best among the single-staining dyes for its ability to stain dead cysts brightly and its relatively slow decay rate of visible light emission following DNA binding. SYTO-9 staining was correlated to animal infectivity. A Live/Dead BacLight was found to be the better of 2 double-staining viability kits tested. Logarithmic survival ratios based on SYTO-9 and Live/Dead BacLight were compared to excystation and infectivity results for G. muris cysts exposed to ozone or free chlorine. The results indicate that SYTO-9 and Live/Dead BacLight staining is stable following treatment of cysts with chemical disinfectants.

Animals↗

PubMedDate not supplied

Evaluation of calcofluor white stain for detection of Pneumocystis carinii.

A rapid calcofluor white (CFW) stain for detecting Pneumocystis carinii was evaluated prospectively. Eighty-nine bronchoalveolar lavage (BAL) specimens, 21 open-lung biopsy (OLB) tissues, 2 induced sputums, 1 expectorated sputum, 2 tracheal secretions, and 1 bronchial secretion from 102 patients were examined for P. carinii cysts by both the CFW stain and a modified methenamine silver (MS) stain. Twenty episodes of P. carinii pneumonia were detected: 19 of these episodes were detected by CFW stain, and 16 of those episodes were detected by MS stain. Discrepancies between the two staining methods were resolved by review of the clinical histories and, in one case, by testing an OLB specimen. Neither staining procedure gave false-positive results with any specimen. More cysts were detected in CFW-stained specimens than in MS-stained specimens (p = 0.05). CFW stain is a simple, rapid, and inexpensive method for detecting P. carinii in clinical specimens and is at least as sensitive as MS stain.

Adolescent↗

PubMed1990

Negative staining of proteins.

Negative staining, some closely related alternative preparation techniques and radiation stability are considered. An attempt is made to clarify the mechanism of action and ultimate resolution limit of negative staining. The results of electron diffraction investigation of thermitase microcrystals embedded in glucose and glucose + stains are presented. It is shown that at doses not exceeding 10 electrons/nm2 electron diffraction from thermitase crystals demonstrate diffraction fields up to 0.2 nm. When adding heavy-atom salts to glucose or using negative staining, the relative intensities of reflections change and electron diffraction patterns for every type of heavy-atom additive (or negative stain) have their specific features. Such characteristic changes of reflection intensities indicate specific interaction of these additives (or stains) with the object. In the case of electron diffraction from the crystals stained using the routine negative staining technique the ordering was preserved down to 0.4-0.5 nm. Increasing the dose up to the normal value results in fading of distant reflections. Thus, negative staining with radiation doses less than the critical one could yield resolution down to 0.4 nm. Yet, the structure may change due to interaction with the stain. Nevertheless, the possibility that such resolution could be obtained for a limited number of objects should not be excluded. Some examples of the application of negative staining for investigation of quaternary and domain structure of proteins (nitrogenase, glutamine synthetase, mitochondrial ATP-synthase, membrane monooxygenase enzymes), tubular and two-dimensional protein crystals (catalase, phosphorylase, HWV protein, hydrogenase), as well as ribosomes and bacteriophages are given in the review.

Animals↗

PubMed2005

Digital staining for multispectral images of pathological tissue specimens based on combined classification of spectral transmittance.

In this study, the digital transformation (digital staining) of the 16-band multispectral image of a hematoxylin and eosin (HE) stained pathological specimen to its Masson's trichrome (MT) stained counterpart is addressed. The digital staining procedure involves the classification of the various H&E-stained tissue components and then the transformation of their transmittance spectra to their equivalent MT-stained transmittance configurations. Combination of transmittance classifiers were designed to classify the various tissue components found in the multispectral images of an HE-stained specimen, e.g. nucleus, cytoplasm, red blood cell (RBC), fibrosis, etc.; while pseudo-inverse method was used to obtain the transformation matrices that would translate the transmittance spectra of the classified HE-stained multispectral pixels to their MT-stained configurations. To generate the digitally stained image, weighting factors, which were based on the classifiers beliefs, were introduced to the generated transformation matrices. Initial results of our experiments on liver specimens show the viability of multispectral imaging (MSI) to implement a digital staining framework in the pathological context.

Diagnosis, Computer-Assisted↗

PubMed2005

An experimental method for testing novel retinal vital stains.

There is uncertainty surrounding the safety of the vital stains currently used to assist macular surgery, and there may be other agents that are more suitable. This study aimed to validate a method of screening retinal vital stains for their potential surgical utility. Bovine retina was exposed to test agents at a range of concentrations. Masked observers determined the minimum dye concentration that reliably stained the retina, defined as the minimum visible concentration (MVC). Computer image analysis (CIE94 colour difference equation) was used to estimate the magnitude of the colour difference between stained and unstained retina. Agents that had favourable staining characteristics underwent safety testing using a retinal pigment epithelium and glial cell culture model. Cells were exposed to each agent and viability was assessed with a mitochondrial enzyme (MTT) assay, and fluorescent live-dead probe (ethidium homodimer-1/calcein-AM). Frozen sections were used to determine which retinal layers were stained. Techniques were tested on the following agents: alcian blue; diethyloxadicarbocyanine; Evan's blue; fast green; fluorescein; Janus green; methylene blue; naphthol green; neutral red; procian (reactive) yellow; rose bengal; and trypan blue. For most dyes, the results of image analysis showed that colour differences increased linearly with dye concentration, although some displayed a more exponential relationship. Five agents showed favourable staining characteristics: Evan's blue, rose bengal, naphthol green, neutral red, and trypan blue (MVC 0.02, 0.01, 0.1, 0.002, 0.01%, respectively). Safety testing of these five agents did not show toxicity, except in glial cells exposed to rose bengal. Relative to the negative control (saline), these showed a 48% reduction in viability using the MTT assay (p<0.001; t=4.71; CI 30-75%), and qualitative damage on fluorescence microscopy. Frozen sections showed that some agents produced diffuse staining of all retinal layers, others produced selective inner retinal staining. There are thousands of biological stains available and many of these may be more effective or safer than those currently used for retinal surgery. This study provides a means of screening potentially useful vital stains.

Animals↗

PubMed2002

Carmine-picroindigocarmine: an alternative multiple staining method.

Carmine-picroindigocarmine, a multiple staining method, was developed at the beginning of the previous century by Professor Harry Kull at the University of Tartu. The stain, combining copper carmine, picric acid and indigocarmine gives bright and colourful results. Nuclear structures are stained in red, cytoplasm in varying shades from yellow to green, collagen fibres in blue, the matrix of hyaline cartilage in greyish-blue, muscle tissues from brownish-red to brownish-green, erythrocytes in yellow. Squamous epithelia are stained in red with horney layers in dark red, nail plate and hairs are stained in bright yellow. The carmine-picroindigocarmine staining is stable, which allows for the combining of additional dyes without interfering with the main colouring. The combination of carmine-picroindigocarmine staining with resorcin-fuchsine in principle maintained the colouring of Kull's original method with additional staining of elastic fibres in violet colour that clearly differentiated them from blue stained collagen fibres. The described multiple staining gives an original colourful and aesthetic result, providing an alternative to other multiple staining methods.

Bronchi↗

PubMedDate not supplied

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.

Eosine Yellowish-(YS)↗

PubMed1996

Characteristics of corneal staining in hydrogel contact lens wearers.

Corneal fluorescein staining is widely used in clinical practice and research, but little information exists on the distribution of staining in a large group of asymptomatic contact lens wearers. This cross-sectional study took place at 3 centers, and investigated the pattern of corneal fluorescein staining in both eyes of 98 hydrogel contact lens wearers. We also investigated the strategies used by three experienced clinicians to grade corneal staining. Overall corneal staining was graded using a scale from 0 to 4 in one-half steps, and five corneal zones, superior, inferior, nasal, temporal, and central, were also graded. The average overall staining grade for both eyes of our subjects was 0.50, with an average of 0.57 for the right eye and 0.44 for the left. This difference between the eyes was statistically significant (p = 0.011). In addition, a comparison of the zones within each eye showed a significant difference (p = 0.0001) among the zones. Corneal staining between the two eyes was also positively and significantly correlated (r = 0.58; p = 0.0001). Grading strategies among clinical investigators were significantly different (p = 0.0001), indicating a potential source of bias in multi-centered studies. The difference in corneal staining between the eyes may represent a source of systematic bias, and could be due to grading the right eye before the left. The correlation in corneal staining between the eyes indicates that the two eyes of a subject cannot serve as independent data points. One-third of the subjects who participated in this study had notable corneal staining. This finding underlines the importance of regularly checking corneal staining in clinical practice.

Contact Lenses↗

PubMed1993

The adsorption staining technique applied to isolated premessenger ribonucleoprotein particles: a comparison with conventional techniques using electron microscope tomography.

A specific type of premessenger RNP particle, Balbiani ring granules from the dipteran Chironomus tentans, was biochemically isolated and visualized in three dimensions with electron microscope tomography. The particles were prepared for electron microscopy in three different ways: positively stained, negatively stained and adsorption-stained (embedded in polyvinyl alcohol, PVA, and concomitantly stained). The results were compared with those obtained for RNP particles studied in situ in ultrathin sections of plastic-embedded cells. The positively stained particles were compacted and heavily deformed with little or no internal structure. The negatively stained and the adsorption-stained particles were well preserved; the outer contours and the central cavities of the particles were outlined. The internal structure, i.e. the folded 7-nm elementary fibre, could not be recognized in the negatively stained particles. In the adsorption-stained particles, however, the fibre was discernable, although not quite as distinctly demarcated as in the plastic-embedded samples. We conclude that embedding in PVA with concomitant staining with uranyl acetate is a rapid method to obtain both good preservation and staining of isolated RNP particles. The PVA-embedded particles were also found to be sufficiently resistant to irradiation to permit a comprehensive tilt-series to be taken for electron microscope tomography.

Adsorption↗