Scintillation fluid

Cells are maintained in suitable medium and starved of serum, if required. Agonist/inhibitors are added for 30 min first, then [35S]Met (5 iCi/ml medium) is added for 60 to 120 min. Cells are then lysed and cleared lysates (equal amounts of proteins) are loaded onto 3 MM papers. After the samples have been allowed to soak into the paper, the papers are boiled in 5% (w/v) TCA three times (2 min each time), rinsed with 100% ethanol, and oven dried. [35S]methionine incorporation is measured in 3 ml of scintillation fluid. 

A small portion (0.2 ml) of the supemate is removed and placed in a scintillation vial containing 0.5 ml of distilled water and 5 ml of scintilation fluid,... 

After the pellet is dissolved in water add 3 ml of scintillation fluid to produce a clear solution, and... 

The scintillation fluid is added and the solutions are counted for 10 minutes in Scintillation Counter. ... 

Uptake and Elimination. Six mussels were placed into 600 ml beakers containing 300 ml aerated Instant 0ceanR (10-12°C) for a 1/2 hr acclimation period before addition of antipyrine (3 pCi/ beaker) (or other test compound). Aliquots (1.0 ml) were taken at intervals and placed directly into scintillation fluid (10 ml, 3a70B, Research Products International Corp., Elk Grove Village,... 

The samples are vortexed again, and an aliquot is taken and used to determine the total [ H] acetylcholine present ([Lq]) by counting for [ H] after addition of scintillation fluid. 

The filters are dried under a heat lamp, transferred to scintillation vials and counted for radioactivity after addition of 5 ml of scintillation fluid. This is used to estimate the amount of bound radioligand. 

The efficiency of the cDNA probe synthesis is determined by scintillation counting of the flowthroughs from steps 9 and 11 5pL of each flowthrough are added to 5mL scintillation fluid in separate scintillation counter vials. The samples are counted on the P-channel, and the obtained counts are multiplied by a dilution factor of 20. Probes should yield a total of 5-25 x 10 cpm see Note 32). 

The mixture is incubated for 4 min at 85 C to complete the competition for receptor sites and centrifuged. The supernatant is discarded, the pellet is washed gently so as not to disturb the pellet. The pellet is resuspended in water and scintillation fluid is added. For quantitative work, the sample is counted for 5 min, for screening purposes 1 min. 

From Dahlquist et a/. with permission of the authors and the American Chemical Society. Reaction was permitted to proceed to 2-5% completion and was then adjusted with acid or base to pH 5.6. The mixture was extracted with ether, and the extract was washed twice with saturated NaCI, then dried with anhydrous sodium sulfate, and added to scintillation fluid for counting of radiolabeled phenol. 

The filters are removed and transferred into 5 ml scintillation fluid. 

Salicylic acid and Its metabolite were separated by two methods. The first was thin layer chromatography on cellulose with BAW solvent as for the In vivo metabolism studies. A quicker separation was achieved with a polyamide column. The entire 400 pL from an individual assay was placed on top of a 0.8 x 2.0 cm column packed with Polyamide-6 (Accurate Chemical and Scientific Corp.). The salicylic acid metabolite was eluted with 6 mL water but salicylic acid was retained. 3a70B scintillation fluid (Research Product International Corp.) was used to determine the radioactive content of the entire 6 mL of eluant. Separation of salicylic acid and its metabolite by polyamide column chromatography was verified by thin layer chromatography. 

RadAway will not produce excellent results (>90% binding) for lipids or solutions with visible suspended solids and is not recommended for scintillation fluids or solutions containing bleach. 

Recently Schutte and Koenders (44) described a method engineered to overcome problems associated with trapping methods and combustion methods. In their design, the column effluent merges with scintillation fluid and is transported to the counter. As a result of this design (a) Sensitivity of 0.2 nCi was attainable (b) resolution was not lost (c) 1 C and 3H could be distinguished. 

Thymidine Uptake Studies. Tritiated thymidine (52 mCi/ anole 0.1 /rCi/ml) was added to cells for 60 min at 37°C. Cells were then washed with PBS, incubated at 4°C for 15 min in the presence of ice cold 5% TCA, rinsed with TCA and scraped from the flasks with a rubber policeman. Cells were again washed with PBS, and solubilized in 0.1N NaOH overnight. Aliquots were assayed for protein (62) and radioactivity (scintillation fluid 100 ml Biosolve (Beckman, Fullerton, CA.), 7g of PPO and 0.6 g of POPOP per liter of toluene). 

In a passive detector developed by the National Radiological Protection Board (Wrixon et al., 1988), the etched pits in the detectors are filled with scintillator fluid. After exposure to radon, the detector is irradiated with an alpha source, and the resulting scintillations counted with a photo-multiplier tube. In this way, track density over 1 cm2 of detector can be measured in a few seconds. Passive detectors used in the UK National Survey were sensitive down to 20 kBq m-3 h of accumulated exposure, equivalent to a radon concentration of 5 Bq m-3 measured over 4000 h exposure. 

Membrane vesicle suspension should be thawed quickly at 37 °C and stored on ice for about 40 min before use. The transport assay is initiated at 37 °C by addition of membrane suspension (30 pg protein) to the transport assay mixture (110 pi). 20 pi aliquots are removed after 30 or 60 sec intervals, immediately diluted with 1 ml of ice-cold incubation buffer and immediately filtered through nitrocellulose membrane using the vacuum of the filtration apparatus (200 mbar). Filter membranes are rinsed twice with 5 ml of cold incubation buffer, dried and dissolved in 10 ml scintillation fluid to count for radioactivity. 

Measurement of radioactive decay can also be affected by various components present in, or added to, the scintillation cocktail. These components can cause quenching that is, they can decrease the efficiency of the scintillation process. Scintillation counting provides data in counts per minute (cpm). Quenching dictates that the counts per minute detected is less than the actual decay rate, or disintegrations per minute (dpm). Almost every sample encountered experimentally is quenched to some degree for example, 02 picked up by the scintillation fluid from contact with air serves as a quencher. Therefore, researchers frequently count an additional sample containing a standard of known de-... 

Incomplete solubility and associated point quenching constitute a major problem in scintillation counting. Efficient scintillation counting requires that the sample be fully soluble in the excitable organic solvents of the scintillation fluid. However, biological systems, which are usually aqueous systems or assays, frequently contain water or hydrophilic molecules that will not dissolve in standard toluene-based scintillation cocktails. 

A second approach has been developed, in part because of difficulties encountered in disposing of scintillation fluids containing naphthalene, which is insoluble in water. In this approach, solubilizer or detergents are added to the standard toluene-based scintillation cocktails. Hyamine hydroxide, NCS, Soluene-100,... 

Survey your hands and all work areas with a Geiger-Miiller meter after each use of isotopes that emit y rays or high-energy /3- particles. After each use of low-energy /T isotopes, such as 3H, sample your hands and work areas with a damp cotton swab and place the swab in a vial containing an appropriate scintillation fluid to screen for possible contamination. Survey the bottoms of your shoes as well, to prevent tracking of radioisotopes around the laboratory. 

H toluene, 14C toluene unknowns 14C toluene standard of known disintegrations per minute per milliliter Volumetric dispensing bottles Scintillation fluid (0.5% PPO in toluene)... 

Using a volumetric dispensing bottle, tip 5 or 10 ml of scintillation fluid into each of two scintillation vials, depending on the size of scintillation vials to be used. Label the bottles A and B at points on the bottle above the fluid level or on the cap. In the same fashion, label the bottles with your name for later identification. 

Scintillation fluid (5 g of 2,5-diphenyloxazole/L-toluene) Scintillation vials Scintillation counter... 

Fill each scintillation vial with enough scintillation fluid to completely cover each paper segment. 

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