Dye Marking Research Articles

Effectiveness of four methods for tagging juveniles of farm‐reared gilthead sea bream, Sparus aurata, L.

This study tested different tags for each size of farm‐reared gilthead sea bream, Sparus aurata L. juveniles, used in enhancement programmes. The following tags were tested: dye mark, fingerling tag, opercular tag, anchor tag A (filament diameter of 1.20 mm) and anchor tag B (0.85 mm). Tag/mark retention, wounds produced by the tag, ease of application, ease of detection and cost were examined. Epidermal dye marks had poor retention and detection rates. Of the tags tested, retention was greater for the anchor tag A (80–100%) after 7 months; anchor tag B showed a retention rate of only 20% after 3 months. The fingerling tag had a retention rate of 27.5% after 12 months. The opercular tag had a retention rate of 44% after 20 days but fractured the opercular bone in many cases. Tag incisions and wounds persisted in some fish for the duration of the experiment.

Thoracoscopic resection of pulmonary nodules after computed tomographic–guided coil labeling

Background. A limiting factor in performing video-assisted thoracic surgery for resection of peripheral solitary pulmonary nodules has been the recognition of the lesion visually. This study reports our clinical experience of injecting a small metallic marker under computed tomographic scan guidance before the operation, allowing localization of the lesion. Methods. A series of 14 patients underwent video-assisted thoracic surgery for removal of 15 pulmonary nodules situated in the outer third of the lung. Before operation, a radiopaque microcoil was injected just behind the lesion and then used to locate, under fluoroscopy, the area to be resected during thoracoscopy. The technique was evaluated for accuracy, reliability, and ease of use. Results. Microcoil labeling of peripheral pulmonary nodules allowed in every case a complete resection and a histologic identification of the lesion. It is more stable and accurate than methylene blue dye marking, and it is as easy to perform as computed tomographic scan–guided biopsy. The incidence of complication was small in spite of our inexperience with the technique. Conclusions. Our experience with microcoil injection shows that it provides consistent and highly accurate marking of pulmonary nodules for video-assisted thoracic surgery, allowing secure resection with a safe margin.

Photoreceptor visual fields, ommatidial array, and receptor axon projections in the polarisation-sensitive dorsal rim area of the cricket compound eye.

We made intracellular recordings from the photoreceptors of the polarisation-sensitive dorsal rim area of the cricket compound eye combined with dye marking. By measuring visual field sizes and optical axes in different parts of the dorsal rim area, we assessed the optical properties of the ommatidia. Due to the large angular sensitivities (median about 20 degrees) and the high sampling frequency (about 1 per degree), the visual fields overlap extensively, such that a given portion of the sky is viewed simultaneously by a large number of ommatidia. By comparing the dye markings in the retina and in the optic lobe, the axon projections of the retinula cells were examined. Receptors R1, R2, R5 and R6 project to the lamina, whereas R7 projects to the medulla. The microvilli orientation of the two projection types differ by 90 degrees indicating the two analyser channels that give antagonistic input to polarisation-sensitive interneurons. Using the retinal marking pattern as an indicator for the quality of the intracellular recordings, the polarisation sensitivity of the photoreceptors was re-examined. The polarisation sensitivity of recordings from dye-coupled cells was much lower (median: 4.5) than that of recordings in which only one cell was marked (median: 9.8), indicating that artefactual electrical coupling between photoreceptors can significantly deteriorate polarisation sensitivity. The physiological value of polarisation sensitivity in the cricket dorsal rim area is thus typically about 10.

Do gravid black flies (Diptera: Simuliidae) oviposit at their natal site?

We present information on an aniline dye marking method for black flies. In the laboratory, adults were sprayed with 2% aqueous solutions of four colors of aniline dyes; brilliant blue and methyl orange gave the best results in longevity trials. In field trials we were able to recapture 1.3% of newly emerged marked and released flies at oviposition sites. Mark–release–recapture experiments were designed to distinguish among three competing models concerning oviposition site selection by gravid female black flies: (1) larval site fidelity (“Do flies return to the site that they experienced as larvae?”), (2) adult site fidelity (“Do flies return to the site that they experienced as adults?”), and (3) no site fidelity (“Do flies oviposit at random, i.e., without regard to adult or larval experience?”). Models 1 and 2 were rejected. There is, however, no reason to reject Model 3, the no site fidelity model. Thus, we conclude that for members of the S. venustum/verecundum complex (i.e., S. rostratum, S. venustum, and S. truncatum) females find an “apparently suitable” waterway in which to oviposit; this may or may not be their natal site.

Intraoperative image-directed dye marking of tumor margins.

The incorporation of interactive image guidance during intracranial tumor surgery offers the possibilities of reduced operative trauma, shorter operation time, greater precision, and an increased understanding of complex anatomy and pathology. A basic weakness with these systems though is that they cannot account for movement of target points due to brain shift by draining of CSF or removal of pathology during the operative procedure. We have developed a stereotactic (frameless) guided injector probe for marking the tumor boundary with dye injection in conjunction with a neuronavigation system. The device consisted of a rigid blunt hollow probe (2 mm dia.) with 4 small side holes at the tip. The catheter is mounted in a holder equipped with 3 LEDs supplying guidance information for the neuronavigation system. A small manual aliquoting pump delivers a measured amount of dye in each track. Isotonic methylene blue was injected in 6 to 8 tracks around the periphery of the tumor as determined by the contract ring in MR scans. The dye was injected using image-directed guidance before resection of the tumor was started (often with the dura intact). Tumor tissue could then be resected until the dye became visible at the tumor boundary. Identification of the dye in the tissue was enhanced with the use of the operating microscope. The 3-dimensional position of the dye track could be determined at the end of tumor resection and compared with its initial position giving a good estimate of local brain shift. The method has proved especially helpful for the resection of large gliomas allowing for a more radical operative result.

Evaluation of marking and tagging methods for genetic studies in carp

A variety of marking and tagging methods were tested on common carp (Cyprinus carpio L.) with the aim of identifying suitable methods for genetic studies in this and other species of carp. Elastomer and Alcian blue dye marking; Cold and Silver nitrate branding; Floy, Fingerling, Carlin disc and visible implant tags; and fin clipping were all tested on a range of sizes of common carp (from mean weights of 10–25 g up to 600–800 g). The branding and tagging methods tested did not give satisfactory retention rates. A combination of elastomer marking and fin clipping was then tested as a method for strain identification in a growth comparison trial on catla (Catla catla Hamilton) and found to be satisfactory for this purpose. Passive integrated transponders (PIT) tags were used to individually identify catla of wild or hatchery origin being grown for use as broodstock. These had almost 100% (98.8%) retention rates, but are expensive compared to most other tagging methods.

Social dominance, prior residence and the acquisition of profitable feeding sites in juvenile Atlantic salmon

Twelve groups of 10 Atlantic salmon Salmo salar parr were screened for dominance in small tanks with a single localized food source using a serial removal method and the top, middle and bottom two fish in each group were classified as high, medium and low social status, respectively. These 72 ranked fish were weighed and formed into four groups or waves, each consisting of six fish of each status category. The fish were given dye marks according to status and wave and were then introduced into an artificial stream in four waves over a 12‐day period. Patterns of settlement/emigration were recorded over a 2‐month period, at the end of which the fish were removed from the stream, identified and reweighed, and daily specific growth rates calculated. Fifty‐one fish established feeding stations in the flume, the remainder emigrating via a downstream trap. The average growth rate over the 2‐month period was 0·87% day−1. Probability of settlement and growth rates following settlement were significantly higher in fish from the earlier waves. In contrast, salmon that were dominant in the initial trial were less likely than subordinate fish to settle in the experimental stream and showed lower postsettlement growth rates. The possibility that fish have individually consistent behavioural profiles that promote success in different competitive environments is discussed with reference to published literature for other groups. A small‐scale trial at the end of the main experiment confirmed the prior residence effect and demonstrated that 1 day was sufficient for such an effect to be evident.

An Evaluation of Some Marking and Trapping Techniques Currently Used in the Study of Anuran Population Dynamics

An Evaluation of Some Marking and Trapping Techniques Currently Used in the Study of Anuran Population Dynamics

Field Trials of Marking Stream Salmonids by Dye Injection and Coded-Wire-Tagging

In the summers of 1993 and 1994, we conducted trials in natural streams to assess the performance of dye marks and full-length coded wire tags (CWTs) when used simultaneously. Brook trout Salvelinus fontinalis and juvenile Atlantic salmon Salmo salar (fork length ≥55 mm) were marked with jet injection of Alcian Blue dye (in the base of the pectoral, pelvic, or caudal fins) and CWT (snout implant). All Alcian Blue marks remained clearly visible for at least 4 weeks. After 8 weeks, 3.1 % of brook trout and 11.4% of Atlantic salmon had lost at least one mark. Although loss rates did not differ significantly among marking locations, 86% of mark losses were from the caudal fin. Marking in pelvic or pectoral fin locations induced high mortality in smaller fish. Dye retention was low for individuals recaptured 10–14 months after injection, presumably because interannual growth caused dilution of the mark. Loss rates of CWT in summer were 2.5% for brook trout and 8.4% for Atlantic salmon; for either species, loss rates did not differ significantly among size-classes. Marked fish did not differ from unmarked fish in growth or condition in summer.

Evaluation of Mark–Release–Recapture Methods for Estimating Forager Population Size of Subterranean Termite (Isoptera: Rhinotermitidae) Colonies

An evaluation of the application of mark–release–recapture models to assess foraging population sizes of subterranean termite colonies is presented. A combination of laboratory and field studies were used to examine the validity of 3 of the underlying assumptions, including persistence of dye marks (Nile Blue A), effect of dye mark on survivorship, and mix of marked and unmarked foragers within the interval between release and recapture in the eastern subterranean termites, Reticulitermes flavipes (Kollar), and R. virginicus (Banks). For both species, the percentage of individuals losing their dye marks when reared on a mixed wood diet was significantly higher than for termites reared on a filter paper diet for 27 d after marking (both treatments permitting limited exercise). There were considerable differences in dye loss between colonies within species. Mortality differences among dyed and undyed termites, although statistically significant, were relatively small and probably do not affect population size estimates derived from mark–release–recapture models. The movement of marked termites between equivalent food resources is unpredictable and contrary to the model assumption of marked termites fully mixing within the population within the interval between recaptures. A simulation is presented to investigate how variables such as dye loss, mortality, and probability of recapture affect accuracy of colony size estimates and their standard errors. The number of recaptures is a dominant factor driving the accuracy of the population estimates generated from these models, but the number of marked termites recaptured in proportion to population size is impossible to assess in field colonies because we do not know the true population size of the foraging colony. Data suggest that validity of mark–release–recapture results may be compromised by the fact that aspects of the biology of subterranean termites violate critical assumptions of the models. Forager population estimates generated by mark–release–recapture studies of subterranean termite colonies must therefore be interpreted with caution.

Effects of recurrent collateral inhibition on Purkinje cell activity in the immature rat cerebellar cortex - an in vivo electrophysiological study

We present here a study on the effects of inhibitory recurrent collaterals of Purkinje cell (PC) axons on the activity of the immature rat cerebellar cortex. Simultaneous extracellular recordings of pairs of PCs were performed in rat pups aged 5–8 days postnatal. Bicuculline was applied to the surface of the cortex in order to functionally antagonize PC recurrent collaterals. At this early developmental stage these are the only inhibitory links in the network. Dye marks from the microelectrode tips and 3D serial-section reconstruction of the structure allowed the exact determination of the distance separating recorded cells and of their respective orientation in the cortex. Standard statistical tests and an informational entropy index were used to calculate levels of cooperativity. By comparing PC activity under control conditions and after bicuculline superfusion it is shown that recurrent collateral inhibition has a structurating effect on the PC activity and that it increases the informational content of the network. Inhibition decreases the activity of the cells by 35% and drastically changes the interspike interval histograms. This leads to a more constrained state of the system. Three types of coupling via recurrent collaterals are present: symmatrical, asymmetrical or non existent. The exact type of coupling follows a simple vicinity rule and strongly influences the cooperativity level between the recorded cells. This cooperativity was also found to be spatially compartmentalized. Several pairs were driven by common inputs via climbing fibers or parallel fibers. Using the predictive value of a theoretical model of this immature structure 16 we propose a complementary explanation of the role of the recurrent collaterals at this stage of development: that of a spatial and temporal filter, specific to each different microzone.

Age validation and movement of freshwater eels (Anguilla dieffenbachiiandA. australis) in a New Zealand pastoral stream

Abstract Forty‐seven eels were captured, tagged, and released into a 420 m lowland section of the pastoral Ahirau Stream, North Island, New Zealand. Eels were marked with jet inoculations of alcian‐blue dye, and plastic tube tags were surgically inserted into the gut cavity. Oxytetracycline hydrochloride was injected intraperitoneally to provide markers on otoliths. Over 3 years, 25 tagged eels were recaptured in the same stream reach, 16 of these within 20 m of their original sites of capture. All recaptured eels retained internal tags which had been incorporated into intestinal mesentary membrane, and 57% of eels recaptured over the first 2 years retained dye marks. Annual length increments, determined from measurement of tagged individuals, were 65 ± 15 mm (mean ± 95% confidence limits) for longfinned eels, and 29±18 mm for shortfinned eels. Translucent zones counted after single fluorescent rings corresponded to the number of years eels had been at liberty. The occurrence of the OTC fluorecence within...

Mutually exclusive expression of alternatively spliced FMRFamide transcripts in identified neuronal systems of the snail Lymnaea

The FMRFamide gene of the snail Lymnaea encodes tetrapeptides (FMRFamide/FLRFamide) and heptapeptides (GDPFLRFamide/SDPFLRFamide) on separate exons. In situ hybridization probes specific to these exons were used to map the expression of the two exons in identified neuronal systems of the CNS. Analysis of more than 200 preparations showed that cytoplasmic expression of mRNA was exclusively of one type, with individual neurons expressing either the tetrapeptide or heptapeptide exon. Of the approximately 340 neurons expressing the two exons, the majority (80%) expressed the tetrapeptide exon. The tetrapeptide exon was more widespread, occurring in neurons from all 11 ganglia of the CNS. The heptapeptide was mainly confined to two ganglia (visceral and right parietal), with a small number of cells in three other ganglia. Mapping studies combined with dye marking of identified neurons showed the presence of the tetrapeptide exon in several behaviorally important networks: heart motoneurons, whole body withdrawal response motoneurons, and probably penis motoneurons as well as giant identified neurons (LP1, RPD1). The heptapeptides were prominent in two main clusters of cells (Bgp and Fgp) together with a smaller number of tetrapeptide-expressing cells.

Enhancement of spanwise coherence in laminar vortex streets

Flow visualization experiments with stripped dye markings show that the vortices in the wake of an impulsively started circular cylinder initially form with random phase along the span. In successive vortex generations, small perturbations disappear within a few cycles, and the eddies align parallel to the cylinder. The spanwise coherence is enhanced if vortex shedding is initiated by trigger plates.

Stratified squamous epithelium in relation to the tympanic membrane: its development and kinetics

The pathways of auditory epithelial migration on the human tympanic membrane and their rate of movement were investigated by Hopkins rod photography of dye markings. The origin of these pathways was determined in both the human and the mouse by studying the development of the stratified squamous epithelium of the tympanic membrane and external auditory meatus from earliest embryonic life to maturity. Two pathways of migration are present. In one, epithelium moves from the region covering the tip of the handle of the malleus upwards to the lateral process and then posterior-superiorly with all dye on the pars flaccida to its posterior superior edge. In the second, dye moves centrifugally and radially outwards from the edges of the handle and pars flaccida regions to the annulus. Rate of movement can be determined approximately only and by reference to anatomical landmarks. The first pathway was traced embryologically to migration possibly commencing in the fundus of the primordial first branchial groove. The second pathway has its source in the growth of the meatal plate. A study of the development of the early meatal plate in the mouse suggests that movement of epithelium over the pars tensa region could be the result of a “pulling” effect of mitotically active cells in a generation center at the edge of the tympanic membrane resulting from negative contact inhibition.

Influence of pre-stocking feeding levels on catchability of rainbow trout, Oncorhynchus mykiss (Walbaum), in a small fishery

. The effects of different pre-stocking feeding levels on the subsequent catchability of rainbow trout, Oncorhynchus mykiss (Walbaum), were investigated in a 12-ha put-and-take fishery. Two sibling batches of Winthrop strain were fed high or low rations of a pelleted diet to produce similar sized, but recently fast- and slow-growing fish. Each batch was given a distinctive dye mark. Anglers’catch records showed that although the 'slow-growing’fish were significantly the more easy to catch during the first week after stocking, over the remaining 5 months of the fishing season there was little difference in catchability between batches. This suggests that previous observations of sustained between-strain differences were not the result of selection of fast- and slow-growing fish per se, but were probably genetically determined.

Cell surface molecules containing N-acetylgalactosamine are associated with basket cells and neurogliaform cells in cat visual cortex

In the cerebral cortex, the plant lectin Vicia villosa (VVA) selectively stains the surfaces of nonpyramidal neurons. This lectin binds specifically to alpha- and beta-linked N-acetylgalactosamine (GalNac). VVA-reactive carbohydrate is highly concentrated in layer 4 of the primary visual cortex of the cat, where it is associated exclusively with GABAergic local circuit neurons. We have studied this neuronal subset with intracellular electrophysiological recording and dye marking to identify the particular cell types expressing surface GalNac. Five different types of local circuit neurons were stained intracellularly (N = 45), but only 2 types, the columnar basket and large neurogliaform cells, were also labeled by the lectin (N = 19/45). Lectin negative types included small basket, chandelier, and large bitufted cells (26/45). Spiny stellate and pyramidal neurons were also lectin negative. Electrophysiological recordings revealed differences in the duration of action potentials in smooth versus spiny stellates but no differences between lectin-positive or -negative types. A biochemical analysis of cortical glycoproteins by SDS-PAGE and lectin blotting revealed multiple bands containing GalNac enriched in membrane fractions. These carbohydrate-containing molecules may be part of a biochemical mechanism for linking together cells with common functional properties.

Evaluation of food dye marking in the study of sandfly behaviour in Baringo District, Kenya

Evaluation of the use of food-dye marking method for the study of sandfly behaviour was carried out in Baringo district, Kenya. A solution of sucrose and food dye was sprayed as bait on the vegetation around a selected source of flies in an animal burrow. The extent of marking and distribution of coloured flies within a radius of 200 m was monitored for four nights. The species caught were Sergentomyia clydei, S. schwetzi, S. antennatus, S. squamipleuris, S. bedfordi, S. africanus S. ingrami and S. affinis and the number of coloured flies was 1017. Feeding on the dyed solution labelled 68.9% of the exiting flies near the burrow and 49.2% of the flies caught at the burrow's entrance. The total number of coloured flies caught at a distance of 50, 100, 150 and 200 m was similar toa linear distribution. The greater numbers of S. clydei caught at 200 m and the smaller numbers of S. squamipleuris and S. ingrami trapped between 50 and 200 m deviated from this pattern. These results are discussed as well as the possible application of the method.

Spheno-occipital synchondrosis--a fluorescence and polarization microscopy study in Cercopithecus aethiops monkeys

On 17 Cercopithecus aethiops monkeys we investigated with the method of the polychromic sequential dye marking system the histomorphology as well as the dynamics of growth and calcification of the spheno-occipital synchondrosis. The age of the animals ranged from change to teeth to late adolescence. The animals had been divided into four different groups: I: late change of teeth; II: young adult; III: fully grown; IV: late adolescence. In the first group the spheno-occipital synchondrosis showed no ossification in the gap which is filled with cartilage. It showed the characteristic structure with a central zone of equally distributed chondrocytes. Adjacent to this we found a zone of proliferation cell hypotrophy and cell degeneration. With increasing age there is decrease of the density of cells (groups II to IV) and after change to teeth the synchondrosis starts to ossify (group II). Due to the ossification the synchondrosis subdivides into different cartilage regions. We found that the closing of the synchondrosis started in the cranial region and progressed toward the caudal region. During this procedure the synchondrosis never ossified completely. Several cartilage regions persisted uncalcified until late adolescence. Interstitial growth of the synchondrosis was found until the end of the change of the teeth (group I). This growth which was always found in a sagittal direction ceased after bony connections had been formed between both poles of the synchondrosis. The sphenoidal and occipital pole of the synchondrosis showed equal growth potential.

Evolutionary modification of cell lineage in the direct-developing sea urchin Heliocidaris erythrogramma

The sea urchin Heliocidaris erythrogramma undergoes direct development, bypassing the usual echinoid pluteus larva. We present an analysis of cell lineage in H. erythrogramma as part of a definition of the mechanistic basis for this evolutionary change in developmental mode. Microinjection of fluoresceinated tracer dye and surface marking with vital dye are used to follow larval fates of 2-cell, 8-cell, and 16-cell blastomeres, and to examine axial specification. The animal-vegetal axis and adult dorsoventral axis are basically unmodified in H. erythrogramma. Animal cell fates are very similar to those of typically developing species; however, vegetal cell fates in H. erythrogramma are substantially altered. Radial differences exist among vegetal blastomere fates in the 8-cell embryo: dorsal vegetal blastomeres contribute proportionately more descendants to ectodermal and fewer to mesodermal fates, while ventral vegetal blastomeres have a complementary bias in fates. In addition, vegetal cell fates are more variable than in typical developers. There are no cells in H. erythrogramma with fates comparable to those of the micromeres and macromeres of typically developing echinoids. Instead, all vegetal cells in the 16-cell embryo can contribute progeny to ectoderm and gut. Alterations have thus arisen in cleavage patterns and timing of cell lineage partitioning during the evolution of direct development in H. erythrogramma.