Filter paper disc incubation

Electrolyte leakage. Tissue discs, prepared from potato tubers as described above, were incubated for 16 h at 25°C between wet filter papers. After incubation, the discs were shaken in 20 ml H2O for another 60 min. One ml of this extract was diluted 30-fold with water, and subjected to conductivity measurements using a HI 8788 apparatus (Hanna Instruments). An increase in conductivity indicates a leakage of electrolytes through lesions in the cell wall caused by enzyme action. Control samples were not incubated, they were shaken in water only. 

Earthworm, Eisenia fetida 32 mg/kg DW soil for 56 days 53.3 (32.5-186.0) mg/kg DW soil for 56 days 210 mg/kg DW soil for 56 days 555 (460-678) mg/kg DW soil for 56 days 683 (570-812) mg/kg DW soil for 14 days Earthworm, Eisenia fetida andrei Adults held in soil containing as much as 300 mg Cu/kg DW for 3 weeks resultant cocoons incubated in uncontaminated soil for 5 weeks to assess hatchability Earthworm, Lumbricus rubellus 100-150 mg/kg DW soil 150-300 mg/kg DW soil >300 mg/kg DW soil 1000 mg/kg DW soil for 6 weeks Earthworm, Lumbricus terrestris Exposed for 5 days to filter paper disc containing 0.5, 1,2, 4, or 8 pg Cu/cm ... 

Mention should be made of a simple enzymatic screening method whereby individuals with very low ceruloplasmin blood levels can be detected (A4). The method employs a filter paper disc (Whatman No. 3). A drop of 0.4% p-phenylenediamine in acetate buffer of pH 5.2,1.0 M, and a drop of plasma are placed on the filter paper in succession. The discs are then wrapped in a polyethylene sheet, placed in a tray, and incubated at 37-40°C. A control sample with a borderline low ceruloplasmin level is included among the unknowns. The intensity of the blue color, which develops as the result of ceruloplasmin-catalyzed oxidation of p-phenylenediamine, is noted and compared to that produced by the standard. Convenient kits for the performance of this screening test can be obtained commercially. ... 

We recently surveyed a cross-section of plants from many tropical regions of the world in a search for photosensitizers to further test the above hypothesis. The methods used to test for phototoxic phytochemicals are described in detail elsewhere (22). Briefly, methanolic extracts were spotted onto sterile filter-paper discs and allowed to dry. The dried discs were placed onto replicate nutrient agar plates that had been spread with Ex B/r (a UV resistant bacterium). The plates were incubated in the dark at 3TC for 30 min. Half of the plates were irradiated for 60 min. with eight Sylvania F40BLB UVA lamps (18W m ), while the other half were kept in the dark. All plates were incubated overnight in the dark at 37°C, after which the zones of inhibition surrounding the filter paper discs were measured. 

Petri dishes are prepared with agar medium seeded with Euglena. Filter paper discs are soaked in the solution under test and placed in the seeded agar. After incubation, the zones of stimulation are measured. The method described in the following section is essentially the method of Robbins, Hervey, and Stebbins (33). 

After addition of 5 pCi of l-[ H]-leucine, the cells were further incubated for 30 min at 30°C. Samples of 100 pi of this suspension were put on filter-paper discs which were dipped into 10% trichloroacetic acid and further treated according to the procedure of Mans and Novelli. ... 

The antibacterial activity of the compounds is determined by the disc diffusion method [42]. The bacteria are cultured in nutrient agar medium and used as inoculum for the study. Bacterial cells are swabbed onto nutrient agar medium (prepared from NaCl (5.0 g), peptone (5g), beef extract powder (3g), yeast extract powder (3g), and agar (20 g) in 1000 ml distilled H2O) pH 7.5 0.2 in Petri dishes. The test solutions are prepared in distilled water to a final concentration of 1 %, 2%, and 4% and then appHed to filter paper discs (Whatman No. 4, 5 mm diameter). These discs are placed on the already seeded plates and incubated at 35 2 °C for 24 h. The zones of inhibition around the discs are measured after 24 h. Co-trimoxazole is used as a standard positive control (Table 6.9). 

To determine the efficiency of aminoacylation of [14C]Phe-tRNA, 5 fil aliquots of the aminoacylation mixture are withdrawn before and after the reaction the samples taken from the reaction mixture at the end of the incubation are spotted onto 3-MM paper discs (Schleicher Schuell) and processed by the cold TCA precipitation method, while the sample taken before the reaction is spotted on a paper disc pretreated empty by the same cold TCA procedure. Determination of the radioactivity present on these filters by liquid scintillation counting allows one to calculate the aminoacylation efficiency of the reaction (which, for phenylalanine, should be >2% of total tRNA). The specific activity of the [14C] Phe-tRNA can be determined after one-step purification of Phe-tRNA by BD cellulose chromatography (Gillam et al., 1968), followed by determination of the radioactivity and of the A260. 

Second instar C. puncticollis larvae (11 days old) were obtained by allowing adult mated weevils to oviposit on the roots for 24 hours. The age of the weevils was recorded from the first day of incubation. One larva was placed into each diet burrow with 5 burrows on each Petri dish. The Petri dishes were then covered with a disc of filter paper to absorb excess moisture from the diets and the lids were replaced and the bio-assay left to stand for 10 days. The percentage larval mortality was recorded. 

For incubation each 4 discs vrere transferred to small petri dishes with 1 ml NL II containing the indicated concentration of cold L-Phe (0 - 75 /Ug/ml) and/ or other additions. Incubation was started by adding the same amount of U- C-L-Phe to all variants (0.15 - 0.6 /Ug/ml, total activity 4.4 - 17-6 x 105 dpm). After incubation discs were sucked on filter paper, floated 4 min, on ice water, again sucked on filter paper and aftervards frozen on dry ice. For analysis radioactivity in the TCA-soluble and the TCA-insoluble (protein) fractions as well as radioactivity of alkaloids excreted to the nutrient solution were determined (l, 3). 

Ethylene Release Ethylene release froM excised leaf discs during incubation in sealed flasks was quantified by gas chroMatography. The COx content of the flask headspace was Maintained by inclusion of a centrewell containing filter paper and sodiuM bicarbonate in phosphate buffer. The 00 level was varied by changing the buffer pH and the bicarbonate concentration (9). Headspace (X3 levels were Monitored by infrared gas analysis. 

A rapid test based on ATP/bioluminescence (106) is also available for kidney prescreening. In this test, a paper filter disc is inserted for 30 min into the renal pelvis of the kidney sample. It is then transferred to a cuvette and incubated with a commercially available test culture Bacillus subtilis BGA at 40 C for 3 h in presence of trimethoprim. The amount of ATP is estimated on the basis of the light released after addition of luciferin/luciferinase reagent. In a variation of this test (107), small cubes of kidney tissue are excised and frozen to be placed directly upon four agar plates, each seeded with a different microorganism. The plates are refrigerated for approximately 4 h, and then are incubated overnight at 37 C. 

---