Stain Technic for Nissl's Granules Following Alcohol-Formol Fixation

1. Nerve tissue is fixed 2-4 hrs in a 5% solution of strong formalin in commerical 95% alcohol.

2. If dehydration is perfect, either chloroform or xylol may be used as a clearing agent.

3. A slow method of paraffin infiltration is advisable.

4. Sections should be cut 10-12 microns in thickness.

5. Coplin staining jars should be annealed by placing them on a rack in a pan of cold water, bringing the water to the boiling point, and allowing the jars to stand in boiling water for twenty minutes.

6. One per cent aqueous solutions of either methylene blue or Grübler's Neutral Roth are used as specific stains for Nissl's granules.

7. These stains are heated to boiling in a beaker, the slides are placed in the Coplin jars which are partially submerged in boiling water, and the hot stain poured into the jars. The flame beneath the water bath is turned down and the slides left for 20 minutes.

8. The excess of primary stain is washed off in 25% and 50% alcohol and the slides passed rapidly thru the alcohol series to absolute alcohol, and finally to xylol.

9. When counterstaining is desired, nigrosin in 1% aqueous solution, methyl orange, saturated solution in 50% alcohol, or, a 0.5% solution of eosin in 50% alcohol are recommended. These stains are used cold, and the slides are merely dipped in them after the excess of primary stain has been washed out in 25% and 50% alcohol.

10. If a cold primary stain is desired, a saturated solution of thionin in distilled water, acidified with 1% carbolic acid, will prove specific for the Nissl substance. Sections should be stained 5–10 minutes in thionin, then passed rapidly thru to absolute alcohol, and xylol. The same counterstains may be used as in the hot method.

11. Sections prepared by the hot method show little tendency to fade after ten years use.

12. Excepting neutral red, all the stains used in this technic are carried by the National Aniline and Chemical Company and are satisfactory. Coleman and Bell neutral red may be substituted for Grübler's Neutral Roth with good results.     

土壤种子库与矿业废弃地植被恢复研究I.Leonard瓶-罐装置在土壤种子库检测中的应用

１前言土壤种子库的组成和动态的检测主要有两种方法,其一是把土壤样品铺在垫有沙子（经消毒除去沙子内可能有的种子）的花盆或其它发芽框上,给予合适条件使土样中的种子萌发,记录幼苗数及种类［１～５］;另一种是用物理方法分离土样直接得到种子,检测种子活力和统计...     

A Staining Rack for Handling Cover-Glass Preparations

A porcelain staining rack (Fig. 1) has been devised for handling cover-glass preparations. The design is along the general lines of staining racks for slides commonly sold by commercial houses. It consists of three parallel rods—one at the bottom, two on the sides. These three rods are held together by two end pieces. Each of the rods has twelve slots, thus the rack holds twelve cover glasses. The slots have been arranged to hold the cover glasses some distance apart. Circular cover glasses having a diameter of 22 mm. are most desirable. Each of the two end pieces has a hole near the top for inserting one end of the wire tongs or handle (see Fig. 1) which is used for removing the rack from the stender dish. The staining rack is 40 mm. long and 35 mm. high and is designed, as shown in the figure, to fit the ordinary Stender dishes commonly used in many laboratories.     

A Staining Rack for Handling Cover-Glass Preparations

A porcelain staining rack (Fig. 1) has been devised for handling cover-glass preparations. The design is along the general lines of staining racks for slides commonly sold by commercial houses. It consists of three parallel rods—one at the bottom, two on the sides. These three rods are held together by two end pieces. Each of the rods has twelve slots, thus the rack holds twelve cover glasses. The slots have been arranged to hold the cover glasses some distance apart. Circular cover glasses having a diameter of 22 mm. are most desirable. Each of the two end pieces has a hole near the top for inserting one end of the wire tongs or handle (see Fig. 1) which is used for removing the rack from the stender dish. The staining rack is 40 mm. long and 35 mm. high and is designed, as shown in the figure, to fit the ordinary Stender dishes commonly used in many laboratories.     

Success of a low‐sloping rack for improving downstream passage of silver eels at a hydroelectric plant

相似文献   

Effect of temperature,anaerobiosis, stirring and salt addition on natural fermentation silage of sardine and sardine wastes in sugarcane molasses

Conditions for a natural fermentation during ensilage of sardines or their waste in sugarcane molasses (60:40 w/w) were evaluated regarding the effect of temperature (15, 25 and 35 degrees C), anaerobiosis (closed vs. open jars), daily stirring of the mixture, and salt addition to the initial mix at 5% (w/w) level. Successful natural fermentation took place in sardine silages incubated at 25 or 35 degrees C in open jars to reach a pH of 4.4 in about 2 and 1 weeks, respectively. For samples kept at 15 degrees C, the pH decline was very slow and pH did not decrease below 5.5 after one month of incubation. At 25 degrees C, the most favorable conditions for silage of sardine waste in cane molasses, as evidenced by the fastest decline in pH to a stable value of about 4.4, were achieved in closed jars and with daily stirring of the mix. The pH 4.4 was reached in one week with an advance of at least 3 days compared to the other conditions (open jars and closed jars without daily stirring). Addition of salt at 5% (w/w) in the mix before incubation inhibited the fermentation process.     

Long-term storage and regular repeated use of diluted antisera in glass staining jars for increased sensitivity, reproducibility, and convenience of single- and two-color light microscopic immunocytochemistry

A procedure is described for the dilution and storage of antisera in glass staining jars into which whole slides are immersed for incubation during light microscopic neuropeptide immunocytochemistry. Diluted antisera, stored at 4 degrees C and continuously reused, were found to be stable for long periods of time (to date over 3 years), and consistently yielded high quality staining in both single- and two-color immunoperoxidase staining. We found this procedure to be more convenient than conventional incubation procedures, allowing the more rapid processing of large numbers of slides and reducing the loss of slides due to technical errors. The consistency and reproducibility of day to day staining were also improved. The immersion of whole slides into the antisera permitted the use of long incubation times (up to 7 days) without the sections drying out, which in many cases substantially enhanced the sensitivity of the staining obtained. A procedure for two-color immunoperoxidase staining is described using diaminobenzidine for a brown color and alpha-naphthol/pyronin for a red/purple color. We found the alpha-naphthol/pyronin reaction superior to the more commonly used 4-chlornaphthol reaction as a second color. The two-color staining was found useful not only for demonstrating nerve cell bodies stained different colors, but also for staining nerve terminals one color that are around and contacting nerve cell bodies stained another color.     

Heteropteran insects as mosquito predators in water jars in southern Vietnam

Residents of Vietnam living in areas with water shortages and/or poor tap water maintain water storage containers, such as jars, in and around their domiciles in order to store water used in daily life. Although these water jars are known to be important breeding sources of the Aedes mosquito, use of chemical larvicides in such containers is legally prohibited in Vietnam. In this study, we identified the dominant mosquito insect predators in water jars in and around residences located in Tan Chanh, Long An, southern Vietnam. Of 3,646 Heteroptera collected from such jars, Corixidae (Micronecta spp.) and Veliidae (Microvelia spp.) were revealed to be the dominant predators. Polymerase chain reaction (PCR) analysis revealed that 40% of Micronecta and 12% of Veliidae had Aedes aegypti‐positive reactions, indicating that these two dominant Heteroptera are important predators of Ae. aegypti. Our results suggest that aquatic Heteroptera may be an important mosquito control agent in addition to the currently used copepods.     

A simple,inexpensive apparatus for rotating bottles during zooplankton feeding experiments

By using the bearings in the hub of bicycle wheels to support the weight of jars containing zooplankton and experimental food, a small laboratory stirring motor can be used to slowly rotate several kilograms of jars for zooplankton feeding experiments. This apparatus is quickly assembled from easily obtainable parts at almost no cost.     

Sequential release and residual activity of temephos applied as sand granules to water-storage jars for the control of Aedes aegypti larvae (Diptera: Culicidae).

Two long-term experiments were carried out on the release profile and efficacy of temephos 1% GR (sand granules) against Aedes aegypti larvae in water-storage containers. In the first experiment, the efficacy of temephos 1% GR enclosed and tied in a muslin cloth and placed in water at the bottom of 200 L earthen water-storage jars was studied by exposing the packets for four to nine wk in one set ofjars and then transferring them sequentially to new sets ofjars four times successively. Temephos released slowly from the granules, the magnitude of release being adequate in the initial period of two to three wk after treatment. Following this period, the efficacy of the granules increased substantially where 92-100% inhibition of emergence even at the lowest dosage of 1 g/100 L (0.05 mg/L AI) was obtained for about another five mo or longer. On removal of the packets from a given set of jars, the released residues remaining in the jars and water lasted a maximum of one to six wk post-removal depending on the magnitude of prior release into the jars. This experiment provided clear evidence that temephos is released slowly over a long period of time in water-storage jars. In the second experiment, we compared the efficacy of temephos 1% GR at 1 and 10 g (0.05 and 0.5 mg/L AI) per 200-L water in jars painted and unpainted on the inside. The efficacy in the painted jars, although high, was consistently lower than that in the unpainted jars, where 99-100% control of larvae was achieved at both rates for a minimum of five mo after treatment. On the basis of this experimental evidence, it is desirable to study the efficacy of lower dosages of temephos than those currently used in Ae. aegypti control programs. The use of controlled release formulations or sachets that are retrievable during cleaning and washing will be more practical and desirable. Both of these interventions will make the program more cost effective.     

An Anaerobic, Intrachamber Incubator for Growth of Methanosarcina spp. on Methanol-Containing Solid Media

To simplify the incubation of Methanosarcina spp. on solid agar medium, a two-port, manual, rectangular air lock was modified to serve as an anaerobic incubator. In one operation, it is possible to incubate 153 petri plates, the equivalent of 11 standard anaerobic jars, with plating efficiencies identical to those of traditional protocols.     

A comparative study of the development of Eimeria nieschulzi in vitro under aerobic and reducing conditions

Sporozoites of the rat coccidian, Eimeria nieschulzi Dieben, 1924 (Apicomplexa: Eimeriidae), were inoculated onto monolayers of normal rat kidney (NRK) fibroblasts and cultured either under aerobic (5% CO2/95% air) or reducing (desiccator jars modified into candle jars) conditions in RPMI-1640 supplemented with 5% fetal bovine serum, sodium bicarbonate, and antibiotics. Under aerobic conditions, first-generation meronts were observed at 2 days postinoculation (DPI) and, except for individual third-generation meronts that were seen at 5 and 6 DPI, no further development was noted. Under reducing conditions, however, first-generation meronts observed at 2-5 DPI underwent additional development to form second-generation meronts (3-5 DPI), third-generation meronts (3-7 DPI), and a small number of fourth-generation meronts (5-8 DPI). Both second- and third-generation meronts were abnormal, exhibiting gigantism although the merozoites produced appeared normal. The gradual degeneration of cell monolayers under reducing conditions prevented further observations beyond 8 DPI. These results suggest that atmospheric conditions play an important role in the development of E. nieschulzi and maintenance of reducing conditions may be one key to achieving enhanced development of some species of coccidia in vitro.     

Effect of incubation conditions at 55 C on moisture loss from agar plates

The percentage of moisture loss was least from plates incubated in plastic bags, sealed jars, or in a humid chamber. There was a stacking effect noted in plates incubated in plastic bags and sealed jars.     

改良Pereira髓过氧化物酶快速染色法及应用

目的　为了使髓过氧化物酶 (MPO)染色快速准确、安全可靠 ,对Pereira碘化钾MPO染色方法做了进一步改进。方法　采用将碘化钾溶于Wright Giemsa染色液中的新配方 ,使试剂更稳定、保存时间长 ,并简化了操作、缩短了染色时间。结果　此法与Washburn联苯胺法比较 ,两者阳性率十分相近 ,差异无显著性意义 (P >0 0 5 )。结论　改良Pereira髓过氧化物酶法阳性反应标本存放多年不褪色 ,是目前众多碘化钾法中较理想的MPO染色方法之一。     

‘Chillum’ jar,a better technique for screening of rhizobia under summer conditions

Summary In the present investigations ‘Chillum’ jar assembly was found to provide more favourable environmental conditions for rhizobia to nodulate leguminous plants particularly under summer conditions than the usual Leonard jar assembly. When thirty pigeon pea rhizobia isolates were tested for their nodulation efficiency in both Leonard jars as well as ‘Chillum’ jars, it was noticed that there was no nodulation in any of the isolates under Leonard jars whereas all isolates were nodulating well under ‘Chillum’ jars conditions. This was probably due to lowering of temperature in ‘Chillum’ jar caused by rapid evaporation from the outer surface of ‘Chillum’ jar assembly. The maximum temperature recorded in ‘Chillum’ jar was 34°C whereas in Leonard jars it was 46.5°C.     

A Combined Silver and Cholinesterase Method for Studying Exact Relations Between the Pre- And the Postsynaptic Elements at the Frog Neuromuscular Junction

Combination of Karnovsky's cholinesterase staining with silver impregnation of axons (modified Bodian's technique) offers a new means of studying the relation between the pre- and postsynaptic elements in the frog neuromuscular junction. The method can be applied to whole muscles so that synapses of individual superficial muscle fibers which have previously been investigated by electrophysiological techniques can be identified after staining. In this way synaptic activity can be correlated with such synaptic features as number of axon branches, length of the occupied synaptic gutter, axonal sprouts, etc. The distinction between occupied and unoccupied parts of the synaptic gutters is useful when studying reinnervation, regression, or growth of a synapse.     

A combined silver and cholinesterase method for studying exact relations between the pre- and the postsynaptic elements at the frog neuromuscular junction.

Combination of Karnovsky's cholinesterase staining with silver impregnation of axons (modified Bodian's technique) offers a new means of studying the relation between the pre- and postsynaptic elements in the frog neuromuscular junction. The method can be applied to whole muscles so that synapses of individual superficial muscle fibers which have previously been investigated by electrophysiological techniques can be identified after staining. In this way synaptic activity can be correlated with such synaptic features as number of axon branches, length of the occupied synaptic gutter, axonal sprouts, etc. The distinction between occupied and unoccupied parts of the synaptic gutters is useful when studying reinnervation, regression, or growth of a synapse.     

Differences in population parameters of twoTribolium castaneum strains in environments of different shapes

1. Survival and duration of development of two Tribolium castaneum strains were not the same when reared in environments of different shapes under the same external conditions.
2. The bb strain survived better in jars than vials. The ++ strain survived better in vials than in jars of type 3.
3. Both strains (as well as their hybrid (+b)) developed faster in vials (type 1) than in shallow jars (type 2). Both strains developed faster in jars of type 3 than in vials, at the low density. At the high density, the strains reacted differently: ++ developed faster in vials, but bb developed faster in jars.
4. Cultures of “selected” strains followed the population trends they exhibited in the selection experiments from which they came.
5. In both densities and in all environments, mean individual weight of ++ adults was always higher then that of bb, suggesting that this character is, at least in part, genetically determined.
6. The differences in survival and developmental period in environments of different shapes may be due to microclimatic differences between environments and to changes produced in them by the developing immatures.

     

A rapid immunogold-silver staining for detection of bromodeoxyuridine in large numbers of plastic sections, using microwave irradiation

Summary A rapid and convenient method for the large scale, immunogold-silver staining (IGSS) of bromodeoxyuridine (BrdU) incorporated by S phase cells, by means of a monoclonal antibody (anti-BrdU) is described. Nineteen slides at a time can be incubated with the antibodies and the protein A-gold (PAG) in staining jars. The antibody and protein A-gold solutions could be used at least five times to incubate new batches of slides. The incubation times with these solutions were shortened by means of microwave irradiation. In this way 200 slides carrying at least 800 sections could be easily processed under the same conditions in one day, using 1.25ml neat antibody solutions of anti-BrdU and rabbit anti-mouse.For light microscopy bothpplastic embedding systems: methylmethacrylate (MMA) and glycolmethacrylate (GMA) can be stained with this technique. The MMA sections, of which the plastic has to be removed before the IGSS, has the advantage of a stronger labelling intensity. The GMA plastic, which contains a cross-linking, agent cannot be removed and consequently for GMA sections it is necessary to incubate the sections with a proteolytic enzyme (trypsin) before the IGSS, to reexpose the antigenic binding sides. However, the GMA sections can be allowed to air dry during the IGSS without negative effects on the morphology. This makes it possible to perform the antibody and the PAG-incubating steps on one day and to finish the IGSS the next day. In this way twice as many GMA slides can be incubated with the same antibody and PAG solutions than with MMA slides.In both plastic embedding systems the intensity of the BrdU labelling was found to be stronger in Carnoy's than in Bouin's fixed sections.     

A novel hematoxylin and eosin stain assay for detection of the parasitic dinoflagellate Amoebophrya

The parasitic dinoflagellate Amoebophrya infects broad range of marine organisms. Particularly, Amoebophrya infections in planktonic dinoflagellates can prevent or delay the formation of algal blooms, and recycle undergrazed planktonic dinoflagellates back to the microbial loop by disrupting host cells. Its ecological significance was gradually recognized along with the discovery of its enormous molecular diversity in oceanic and coastal ecosystems. Thus, we developed a reliable, easily accessible and less time-consuming assay, to detect and assess Amoebophrya infections in planktonic dinoflagellates. The modified hematoxylin and eosin staining assay provided reliable diagnosis of Amoebophrya infection by identifying the characteristic “beehive” of the multinucleate trophonts. After staining, the typical multinucleate “beehive” is evidently distinguishable from the compact nuclei of uninfected host cells. The modified hematoxylin and eosin (H & E) staining assay is easy to use, that can be routinely performed within 3 h (up to 20 samples/batch) using general laboratory equipment, supplies and chemical reagents. The produced slides with agar-embedded dinoflagellate cells can be stored for several months or even years in a dry place without noticeable loss in quality of staining. With suitable calculation, the modified H & E assay can be applied to assess the prevalence of Amoebophrya infection in planktonic dinoflagellates. This efficient and powerful assay will facilitate the investigation on the ecological roles of Amoebophryidae in coastal and oceanic ecosystem.