Volume tissue sample

Add enough perchloric arid to cover the tissue sample. Add the same volume of perchloric acid to the standard saffl es.ctf DNA. 

Regular, routine sample recovery measurements can be made by using the method of standard addition. The matrix is spiked with the analytes in a small volume of solvent at a level which is 50%, 100%, 150%, and 200% above the estimated level in the sample. A number of independent replicates should be made at each level. Provided that sufficient material is available the sample can be analyzed prior to spiking. In case of limited size (e. g., small tissue samples) a number of samples may be pooled and homogenized for such recovery experiments. 

The tissue is homogenised with Chloroform Methanol (2 1, v/v) to a final volume 20 times the volume of the tissue sample (1 g in 20 ml of solvent mixture). 

Mix A solubilized, aqueous tissue sample with an equal volume of ice-cold Soln. A, and then add ice-cold Soln. B up to 3.0 ml. The mixture is left for 10 min in an ice bath. After this centrifuge, leave the acidic solution at 0 °C for 10 min with 4000 x g. Wash the pellet three times by resuspension in ice-cold Soln. C and further centrifugation. 

Aqueous samples are extracted with methylene chloride. A 1-L volume of sample is repeatedly extracted in a separatory funnel. The methylene chloride extract is exchanged to hexane during concentration to a volume of 1 mL. Nonaqueous samples, such as soils, sediments, sludges, fly ash, and tissues may be extracted by Soxhlett extraction or sonication. Methylene chloride, toluene, hexane, or a combination of these solvents may be used for extraction. Sludges containing 1% or more solids should be filtered. The aqueous filtrates and the solid residues are extracted separately. They are then combined prior to cleanup and analysis. 

Microdialysis sampling has been applied to numerous tissues, especially the brain since the brain is sensitive to alterations in volume and ionic composition. Ultrafiltration has been primarily used for peripheral tissue sampling from subcutaneous tissue since the removal of fluid from the brain is believed to cause alterations in brain chemistry.2 For basic research use, microdialysis sampling devices are typically called microdialysis probes. For clinical studies, the device is called a microdialysis sampling catheter since in clinical medicine a catheter is defined as a small tube that can be implanted. 

Figure 6.4 Tissue parameter effects on microdialysis sampling extraction efficiency(EE) using the published Bungay et al. model.42 (a) Parameters in graph (a) denote changes with diffusion coefficients combined with the following parameters probe length, L, 0.3 cm inner radius, ri 0.021 cm outer radius, r0,0.025 cm cannula radius, rcannuia, 0.0175 cm tissue sample volume fraction, ( )s, 0.2 membrane diffusion coefficient, Dm, 0.2Dd tissue sample diffusion coefficient, Ds = Dd/2.25 K,.p 1.0 min-1, Km 0.1 min-1. In vitro simulation sets the external tissue resistance, Rc to a value of zero, (b) Variation in tissue sample volume fraction, <(is, with all parameters used. Dd —5 x 10 6 cm2/s. (c) Variation in sum of external rate constants. All other parameters are as in (a) with Dd— 5 x 10-6 cm2/s.

A two mL capacity polypropylene container with sealable screw-top lid and V shaped bottom. Ideal for storing dried organisms (e.g., amphipods) and water samples and good for digesting small tissue samples because small acid volumes remain in contact with tissue samples. Volume 1(12). 

In contrast to nucleic acids where polymerase chain reaction (PCR) allows the investigation of single cells, no amplification technology is available at the protein level. As such, proteomic studies require relatively large sample volume. Because of this limitation, it is not a surprising that most proteomic studies have been performed on whole tissue samples. This approach in cancer research is, however, unfortunate because cellular heterogeneity within a biopsy or tissue sample will likely impair the quality and reproducibility of information generated. 

Blood samples are taken during the study or at autopsy. Gonadal steroid hormones are determined by specific immunoassays requiring very small sample volumes, extraction procedures are no longer needed. Steroid receptors can be determined in tissue samples storage at autopsy, for later decision on necessary investigations. 

The sensitivity quoted above is for a dry sample and this is greatly reduced by the presence of water. This raises difficulties in the study of samples such as tissue slices, blood, or cell cultures in in vivo conditions. Specially shaped sample holders have to be used and the sensitivity is reduced by a factor of about 100 (allowing for the smaller volume of sample which can be introduced into the spectrometer). A way round this difficulty is to freeze the sample (which restores the full sensitivity), and then to record the spectrum at a low temperature, usually that of liquid nitrogen (77°K). 

Liquid nitrogen is used to store biological tissue samples at low temperatures, o Increased pressure allows a larger mass of propane to fit into a smaller volume for easier transport. Propane is sold as LP (liquid propane) for this reason, although it is actually burned for fuel as a gas. 

Dionex Technical Note 342, Determination of PCBs in Large-Volume Fish Tissue Samples Using Accelerated Solvent Extraction (ASE). 

This method is based on the quantitation of total amino acids following acid hydrolysis of proteins present in tissue samples. Microtiter plates are used in this assay. Tissue samples (10 pg) are first hydrolyzed in 500 pL of 6 M HC1 at 100 °C for 24 h to liberate ammonium. The samples are then lyophilized (chilled and evaporated), and the residue, containing ammonium chloride, is dissolved in a known volume of water. 

Halothane in blood was determined by extraction with an equal volume of carbon disulfide and measurement of the absorption at 7,90 microns in a 10 mm. microcell (80). The method was not suitable for tissue samples. 

A 5—500 pi aliquot of the aqueous sample is precisely selected (Fig. 1.3a) and introduced into the flow manifold. This is especially relevant for the analysis of biological fluids, cell tissues, blood sera, dew and other volume-limited samples, as well as for in vivo assays. Moreover, sampling strategies relying on mini-probes become more practical. In spite of the low sample volume, reliable results are obtained even for very low analyte concentrations. 

---