一种用于食草动物粪便显微组织分析的临时装片新技术

粪便显微组织分析法是研究食草动物食性的主要方法,其常规装片技术需要使用Hoyer's 装片介质对植物碎片进行封片,而Hoyer's 封片液的粘性易导致植物碎片在装片过程中发生卷曲和重叠,影响植物碎片的识别效果。本文提出的新装片技术采用没有粘性的饱和NaCl 溶液代替Hoyer's 装片介质,结合特定的定量取样方法和装片程序,可以有效地减少植物碎片的卷曲率和重叠率。对比试验显示,新装片技术可使植物碎片卷曲率从10.4% 下降至3.8%,重叠率从25% 下降至8.1% ,说明新装片技术在减少植物碎片卷曲和重叠方面明显优于常规装片方法。     

Analysis of the Changes In the Proportion of Clustered Labelled Cells In Epidermis

Abstract. A new cell kinetic approach is presented from which the duration of the S and G₂+ M phases can be estimated. the technique involves an analysis of the spatial distribution of labelled cells in sections or sheets of epithelium (i.e. an analysis of clustered labelled cells). the technique is largely independent of the absolute number of labelled cells and hence is not influenced by factors which affect the absolute number of labelled cells. the technique is described and experimental data from dorsal murine skin are presented. the technique has also been simulated mathematically so that the phase durations and their variances could be estimated. the advantages of the technique are: (1) it is technically simple; (2) it provides at least two independent estimates of the phase durations; (3) unlabelled cells need not be counted (compare with LI or PLM analysis); (4) it is independent of variations in the absolute yield of labelled cells, and (5) it is applicable if the LI is low and the S phase is short (where the PLM technique tends to fail).     

Improving the precision and accuracy for estimating energy expenditure using the heart rate method

The "heart rate technique" is commonly used to estimate the rate of oxygen consumption (a proxy for energy expenditure) of free-ranging animals. However, a major limitation of this technique is that interindividual variability in the relationship between heart rate (f(H)) and rate of oxygen consumption (Vo2) generates large errors of estimation when the technique is applied to individual free-ranging animals. In this study, we present a new analysis technique that takes advantage of the observation that the f(H) or Vo2 relationships between individuals are frequently parallel and differ only in elevation. This technique offers superior accuracy and precision of Vo2 estimates, reducing the coefficient of variability from 18% to 9% for individual animals in an example application in macaroni penguins. This approach enables application of the heart rate technique to deduce the energetic strategies of individual animals.     

A Technique for Continuous Observation of the Infection Process in Cucumber Anthracnose

A new light microscope preparation technique for high magnification observation of living plant tissue and fungal penetration is described. Agar immersion is used in differential interference contrast microscopy (DIC) instead of coverslips (lens 40) or instead of coverslips and oil (lens 100).
This technique is suitable for
(a) longtime observation of living tissue, because the tissue to be observed remains on the plant, and for
(b) thick and uneven samples, as no coverslips are required.
With this technique it was possible to observe the dynamics of penetration of Colletotrichum lagenarium into epideral cells of cucumber cotyledons for 72 hours.
A time lapse film using this technique is in preparation.     

An improved assay technique for matrone, a mosquito pheromone, and its application in ultrafiltration experiments.

An improved assay technique is presented for the detection of matrone, the mosquito pheromone which prevents insemination in female mosquitoes. This new technique requires 24 hours less to complete. The number of repeat tests necessary is reduced by 50% as compared to the number required utilizing the old technique and the reliability of the test is improved through the visual surveillance of the flight ability and mating activity of the females. Incorporated into this assay technique is the construction and use of a dose (dilution) response curve (calibration line) with a confidence band, establishing a threshold level of matrone activity and making it possible to assess the dilution of matrone in any non-toxic test material. Utilizing this new technique, we demonstrated by ultrafiltration that the molecular weight of matrone is between 50,000 and 100,000.     

Evaluation of gravimetric techniques to estimate the microvascular filtration coefficient

Microvascular permeability to water is characterized by the microvascular filtration coefficient (K(f)). Conventional gravimetric techniques to estimate K(f) rely on data obtained from either transient or steady-state increases in organ weight in response to increases in microvascular pressure. Both techniques result in considerably different estimates and neither account for interstitial fluid storage and lymphatic return. We therefore developed a theoretical framework to evaluate K(f) estimation techniques by 1) comparing conventional techniques to a novel technique that includes effects of interstitial fluid storage and lymphatic return, 2) evaluating the ability of conventional techniques to reproduce K(f) from simulated gravimetric data generated by a realistic interstitial fluid balance model, 3) analyzing new data collected from rat intestine, and 4) analyzing previously reported data. These approaches revealed that the steady-state gravimetric technique yields estimates that are not directly related to K(f) and are in some cases directly proportional to interstitial compliance. However, the transient gravimetric technique yields accurate estimates in some organs, because the typical experimental duration minimizes the effects of interstitial fluid storage and lymphatic return. Furthermore, our analytical framework reveals that the supposed requirement of tying off all draining lymphatic vessels for the transient technique is unnecessary. Finally, our numerical simulations indicate that our comprehensive technique accurately reproduces the value of K(f) in all organs, is not confounded by interstitial storage and lymphatic return, and provides corroboration of the estimate from the transient technique.     

Rapid MR imaging of cryoprotectant permeation in an engineered dermal replacement

Magnetic resonance (MR) imaging is a powerful technique for monitoring the permeation of cryoprotective agents (CPAs) inside tissues. However, the techniques published until now suffer from inherently long imaging times, limiting the application of these techniques to slow diffusion processes and large CPA concentrations. In this study, we present a rapid MR imaging technique based on a CHESS-FLASH scheme combined with Keyhole image acquisition. This technique can image the fast permeation of Me(2)SO solutions into freeze-dried artificial dermal replacements for concentrations down to 10% v/v. Special attention is given to evaluating the technique for quantitative analysis.     

A wavelet-based soft decision technique for screening of patients with congestive heart failure

A wavelet-decomposition with soft decision algorithm is used to estimate an approximate power spectral density (PSD) of R–R intervals (RRI) of ECG data for the purpose of screening of congestive heart failure (CHF) from normal subjects. The ratio of the power in the low-frequency (LF) band to the power in the high-frequency (HF) band of the RRI signal is used as the classification factor. The trial data used for estimating of the classification factor consist of 15 CHF (patient) subjects and 12 normal sinus rhythm (NSR) or simply normal subjects. The performance of the algorithm is then evaluated on test data set, which consists of 17 CHF subjects and 53 NSR subjects. Both trial and test data are drawn from MIT database. The receiver operating characteristics (ROC) is used to determine the threshold value of the classification factor. Results are shown for different wavelets filters. The new technique shows a classification efficiency of 96.30% on trial data and 88.57% on test data. An FFT-based frequency domain screening technique is also implemented and included in this work for the purpose of comparison with the wavelet-based technique. The FFT-based technique shows an efficiency of classification of 99.63% on trial data and 81.42% on test data. The comparison is also done on short-term (5-min) recordings. The wavelet-based soft-decision technique shows also better results than the FFT-based technique.     

Avidin-biotin-immunoglucose oxidase: use in single and double labeling procedures

We have investigated the use of an avidin-biotin-immunoglucose oxidase (AB-GO) technique for single and double antigen localization in conjunction with the avidin-biotin-immunoperoxidase (AB-P) technique in fixed, embedded specimens, using sequential monoclonal and polyclonal antibodies of the same species. The optimal technique for double labeling requires the first antibody to be applied and localized with the AB-P technique using 3,3'-diaminobenzidine (DAB) as the chromogen, followed by an optional elution step and/or incubation with mild detergent (0.01% Triton). The second antigen is localized with the AB-GO technique with nitro blue tetrazolium (NBT) as a chromogen. Effects of antigen concentration, intermediate elution steps, and the relative efficiency of the two methodologies are described.     

Phosphorimetric method to determine Zn-porphyrins in microorganisms and biological liquids

A phosphorimetric technique of direct quantitative determination of Zn-porphyrins in microorganisms and biological liquids is described. The technique is based on the registration of visible light-induced afterglow and phosphorescence of Zn-porphyrins at room temperature under anaerobic conditions. The sensitivity of the technique is not less than 10(-8) M. Certain types of Zn-porphyrins can be identified with respect to the spectra of excitation and radiation of afterglow. The technique has been applied to study the afterglow of microorganisms (E. coli, St. aureus, B. subtilis, B. pioceaneum, M. tuberculosis) and of milk, serum, and urine.     

Nanosecond optical rotatory dispersion spectroscopy: application to photolyzed hemoglobin-CO kinetics.

A standard technique for static optical rotatory dispersion (ORD) measurements is adapted to the measurement of ORD changes on a nanosecond (ns) time scale, giving approximately a million-fold improvement in time-resolution over conventional instrumentation. The technique described here is similar in principle to a technique recently developed for ns time-resolved circular dichroism (TRCD) spectroscopy, although the time-resolved optical rotatory dispersion (TRORD) technique requires fewer optical components. As with static ORD, TRORD measurements may be interpreted by empirical comparisons or may be transformed, via the Kramers-Kronig relations, to more easily interpreted TRCD spectra. TRORD can offer experimental advantages over TRCD in studying kinetic processes effecting changes in the chiral structures of biological molecules. In particular, the wider dispersion of ORD bands compared with the corresponding CD bands means that ORD information may often be obtained outside of absorption bands, a signal-to-noise advantage for multichannel measurements. Demonstration of the technique by its application to ns TRORD and the transform-calculated TRCD of carboxy-hemoglobin (Hb-CO) after laser photolysis is presented.     

The measurement of shear modulus and membrane surface viscosity of RBC membrane with Ektacytometry: a new technique

A new technique is proposed to estimate the shear modulus (mu) and membrane surface viscosity (eta(m)) of red blood cell (RBC). Theoretical formulae for finding these two parameters are first derived based on the force balance on a RBC in a flow field of low viscosity. Different types of Ektacytometry are then used to measure relevant quantities. The obtained values (mu=6.1 x 10(-6)N/m, eta(m)=8.8 x10 (-7)Ns/m for normal RBC) are consistent with those previously found by micropipette technique and in AC electric field. The present technique is, however, much easier to operate and more advantageous in reflecting the average properties of a large quantity of RBCs, and it is much cheaper to be applied in clinical practice than any other method of measuring the two parameters. The sensitivity of the technique is demonstrated by testing RBCs treated with glutaraldehyde of different concentrations. This technique was demonstrated by the flow chamber.     

Application of wound dressing Molndal technique in clean and potentially contamined postoperative wounds--initial comparative study

Because of a possible delayed wound healing, critical colonization and infection of wounds present a problem for surgeons, particularly in patients with compromised immune system or in case where the wound is heavy contaminated or poorly perfused. Molndal technique of wound dressing has proven to be effective in prevention of infection. In our study we wanted to describe the benefits of the application of Molndal technique wound dressing compared to traditional wound dressing technique at potentially contaminated and clean postoperative wounds. We examined postoperative wound after radical excision of pilonidal sinus and after implantation of partial endoprosthesis in hip fracture. Molndal technique consisted of wound dressing with Aquacel Ag - Hydrofiber. Traditional technique was performed using gauze compresses and hypoallergic adhesives. We analyzed the results of 50 patients after radical excision of pilonidal sinus. 25 patients were treated by Molndal technique and 25 patients by the traditional technique of wound dressing. In the group treated by Molndal technique only 1 (4%) patient has revealed a wound infection, proven by positive microbiological examination and suppuration. In the traditional technique group 4 (16%) patients developed wound infection as inflammation and secretion as a sign of superficial infection. In the other group we analyzed the results of 50 patients after implantation of partial endoprosthesis after hip fracture. 20 patients were treated by Molndal technique and 30 patients by the traditional technique of wound dressing. In the group treated by Molndal technique no patient has revealed a wound infection (0%). In the traditional technique group 4 (13%) patients developed wound infection. All complication in both group were superficial incisional surgical infection (according to HPSC). There was no deep incisional surgical site infection or organ/space surgical site infection. Our results are clearly showing that Molndal technique is effective in preventing the postoperative wound infection.     

碱性磷酸酶标A蛋白加强的PAP技术

本文介绍一种碱性磷酸酶标Ａ蛋白加强的ＰＡＰ技术。采用ＰＡＰ技术、碱性磷酸酶标Ａ蛋白（ＰＡＡＰ）技术ＰＡＡＰ和加强的ＰＡＰ（ＰＡＰ－ＰＡＡＰ）技术显示下丘脑室旁核催产素（ＯＴ）能神经元。结果发现,其中使用ＰＡＰ－ＰＡＡＰ技术免疫反应产物的显色最深。此技术的原理可能是,由于Ａ蛋白分子至少有四个位点能与ＩｇＧ分子的Ｆｃ段高亲合性地结合,故在该技术中,先经过ＰＡＰ程序的三步免疫反应并显色后,每个与一抗结合的二抗分子上和每个与二抗结合的ＰＡＰ复合物分子上各暴露一个能与Ａ蛋白分子结合的Ｆｃ段,在随后经过ＰＡＡＰ技术处理时,部分ＰＡＡＰ复合物分子就结合在这些Ｆｃ段上,经显色后,ＰＡＡＰ技术显示的浅紫兰色与ＰＡＰ技术显示的浅棕褐色重叠,变成更深的反差明显的深棕褐色。     

Comparative evaluation of methods for detecting HBsAg in sera

The results of the analysis of 1209 serum samples, made with a view to detecting those containing HBsAg, are presented. This analysis was made by the radioimmunoassay (RIA) on a polyethylene film, by the standard RIA technique with the use of a diagnostic kit obtained from Abbott Laboratories (USA) and by the passive hemagglutination (PHA) test. The RIA film technique was found to have the sensitivity of about 2 ng/ml HBsAg, which is similar to the sensitivity of the kit from Abbott Laboratories and exceeds the sensitivity of the PHA test approximately 50-fold. The percentage of detected HBsAg-positive sera, yielded by analysis with the use of the RIA film technique and the standard RIA technique, is the same. The RIA technique make it possible to detect more positive sera than the PHA test by about 2.5%.     

Mass spectrometric characterization of proteins transferred from polyacrylamide gels to membrane filters

In the 1990s, a technique was developed to transfer proteins from electrophoresis gels onto poly(vinylidene difluoride) (PVDF) membranes, digest the proteins on the membranes with proteases such as trypsin and analyze the resulting peptides on the membranes directly by mass spectrometry to identify the proteins. This technique, based on gel electrophoresis, is particularly useful for analyzing protein isoforms, splicing variants and post-translationally modified proteins. Previously, the low ionization efficiency of peptides immobilized on the membranes often rendered this technique useless. However, this technique has been improved by the use of PVDF membranes with a small pore size, which has enabled highly efficient and effective electroblotting and mass spectrometric analyses. Here, the advantage of this technique is discussed.     

Validation and utility of novel volume reduction technique for determination of parallel conductance

The parallel conductance volume, created by the conductivity of structures surrounding the ventricular blood pool, can be estimated by using a saline dilution technique. This paper examines the use of a novel volume reduction method, during a standard vena caval preload reduction maneuver, as an alternative to the routinely used saline dilution method to calibrate conductance catheter measurements in the left (LV) and right ventricle (RV) of animals and humans. The serial reproducibility of both methods was examined by measurement of percent difference, and by assessing the coefficient of repeatability 1) between two measurements within the same subject, 2) between the two techniques, and 3) interobserver variability. The effect of ventricular size and contractile state on the volume reduction technique was also observed. It was essential to ensure the technique was not affected by inotropic state. The volume reduction technique and saline dilution method were repeated at three different loading states (baseline, 5, and 10 microg x kg(-1) x min(-1) of dobutamine). The coefficient of repeatability between serial measurements was similar for both the volume reduction and saline dilution methods, and good interobserver variability was demonstrated. The volume reduction technique was compared with the saline dilution technique over a large range of ventricular sizes. No significant difference was observed in the RV or LV of adult humans or in the LV of neonatal pigs and children. There was no significant effect on either the saline dilution or the volume reduction technique as the inotropic state increased. In conclusion, the volume reduction technique is neither affected by ventricular size nor contractile state, is repeatable between different observers, and can be used to substitute the saline dilution method when preload reduction of the ventricle is being employed.     

Technique for Estimating Low Numbers of a Bacterial Strain(s) in Soil

A technique is described for obtaining most probable number estimates of the number of cells of a bacterial strain(s) when it is present in low numbers in the soil. The technique is based on the bacteriophage response that is elicited when a known number of bacteriophage for the bacterium of interest is incubated with soil dilutions in a nutrient broth. The technique was evaluated for use with gram-negative bacteria.     

Colour development of immunogold-labelled antibodies for light microscopy

Summary We describe an improvement of the immunogold technique, which is based on the colour development of silver-intensified immunogold signals. This method (referred to as the coloured-SIG technique) was found to be as sensitive as the silver-intensified immunogold method and more sensitive (in two of the three tested systems) than immunoenzymatic procedures, such as the peroxidase/antiperoxidase technique or the avidin-biotin system. The coloured SIG-method results in either a magneta-red or a cyan-blue final reaction product. Therefore, this new improvement of the immunogold technique should be useful in (1) double-staining methods, (2) immunoblot methods and (3) conventional immunostaining methods.Supported by the Robert-Bosch-Foundation, Stuttgart