Spotting procedure

Correct nrocedure for spotting It is recommended to integrate the three stages of spotting 

Examine textiles for excessive dirt, particularly at collars, pockets, sleeves, and trouser legs. If textiles are to be dry cleaned check them particularly for stains originating from food or body secretions. 

If the textiles are to be wet-cleaned check them particularly for greasy stains. Apply a small quantity of brushing agent onto the stained areas and allow to react for 10-20 minutes before loading the cleaning equipment. 

Intensive staining found during the examination of the textiles can be treated with special spotting agents. The stain substance must be identified and related to one of the following categories  

Stains that could not be removed dining dry-cleaning or wet cleaning with machines are subject to post-spotting. Proceed as follows  

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In addition, any stains not removed either by pretreatment or during the wash cycle can be heat-set in conventional systems through accelerated oxidation or denaturing of stain components. Some heat-set stains can never be removed. By avoiding an elevated-temperature drying step, the Micare process eliminates the heat-setting of stains, and minimizes time spent on post-spotting procedures. 

Spencer AB, Earnest GS, Kovein RJ. 1995. In-depth survey report Exposures during the spotting procedure in a commercial dry cleaner at Widmefs Dry Cleaning, Cincinnati, Ohio. U.S. Department of Health and Human Services, Public Health Service, Centers for Disease Control and Prevention, National Institute for Occupational Safety and Health. Report No. ECTB 201-13b. 27 pages. 

It is crucial that the spotting robot used has to be able to pass the walls of the Lab-Tek chamber. The spotting procedure for 384 samples on 50 Lab-Tek chambers in parallel using 8 solid pins typically lasts 6 hours. In order to avoid evaporation of the small sample volumes in the 384 well plate during spotting, it is cooled with an in-house built water-cooled plate. In order to deposit more sample volume, which for some cell types improves transfection efficiencies, we spot repeatedly to the same position. 

Only a small amount of material is needed. It is often helpful to blow gently on the plate as the sample is applied. This helps to keep the spot small by evaporating the solvent before it can spread out on the plate. The smaller the spot formed, the better the separation obtainable. If needed, additional material can be applied to the plate by repeating the spotting procedure. You should repeat the procedure with several small amounts, rather than apply one large amount. The solvent should be allowed to evaporate between applications. If the spot is not small (about 2 mm in diameter), a new plate should be prepared. The capillary pipette may be used several times if it is rinsed between uses. It is repeatedly dipped into a small portion of solvent to rinse it and touched to a paper towel to empty it. 

Baeumlisberger D, Rohmer M, Arrey TN, et al. Simple dual-spotting procedure enhances nLC-MALDIMS/MS analysis of digests with less specific enzymes. J Proteome Res. 2011 10 2889-94. doi 10H021/pr2001644. 

Using radioactive isotopes, an accurate and sensitive method (cf. p. 155) measurement of the total radioactivity of the whole spot or the scanning technique may be carried out the error is less than 1 %, the principal error resulting from the spotting procedure (Fig. 240). 

Quality Audit. Another important responsibiUty of quahty assurance is the audit function. Using the quahty audit as a tool, QA can monitor the operation of the manufacturing faciUty a toU, ie, contract, manufacturer or raw material suppHer to assure that written procedures are in place and that there is documentation to indicate the procedures are being followed. Properly executed audits allow QA to spot potential weakness in the quahty system that could allow errors to occur. Once identified, these weaknesses can then be corrected before they result in nonconformance. 

The second conversion of GS to (GSi)r will be Case 4 of Table 5-10, the two-surface-zone enclosure with computation simphfied by assuming that the direct-view fac tor from any spot to a surface equals the fraction of the whole enclosure that the surface occupies (the speckled-furnace model). This case can be considered an ideahzation of many processing furnaces such as distilling and cracking coil furnaces, with parts of the enclosure tube-covered and part left refrac-toiy. (But the refractory under the tubes is not to be classified as part of the refractory zone.) Again, one starts with substitution into Eq. (5-173) of the terms GSi, GS, and S Si from Table 5-10, Case 4, with all terms first converted to their gray-phis-clear form. To indicate the procedure, one of the components, S Si, wil be formulated. 

Follow proper start-up procedures. Introduce feed to the riser only when the reactor system is adequately heated up. Local cold spots cause coke to build up in the reactor cyclones, the plenum chamber, or the vapor line. 

The introduction of the sample into the adsorbent layer is a critical process in HPTLC. For most quantitative work a platinum-iridium capillary of fixed volume (100 or 200 nL), sealed into a glass support capillary of larger bore, provides a convenient spotting device. The capillary tip is polished to provide a smooth, planar surface of small area (ca 0.05 mm2), which when used with a mechanical applicator minimises damage to the surface of the plate spotting by manual procedures invariably damages the surface. 

Procedure. Pour the developing solvent into the chromatographic tank to a depth of about 0.5 cm and replace the lid. Take a prepared plate and carefully spot 5 pL of each indicator on the origin line (see Section 8.6, under Sample application) using a micropipette. Allow to dry, slide the plate into the tank and develop the chromatogram by the ascending solvent for about 1 h. Remove the plate, mark the solvent front and dry the plate in an oven at 60 °C for about 15 min. Evaluate the R value for each of the indicators using the equation... 

Carefully scrape the separated bromophenol blue spots on to a sheet of clean smooth-surfaced paper using a narrow spatula (this is easier if two grooves are made down to the glass on either side of the spots). Pour the blue powder into a small centrifuge tube, add 2 mL of ethanol, 5 drops of 0.880 ammonia solution, and stir briskly until the dye is completely extracted. Centrifuge and remove the supernatant blue solution from the residual white powder. Repeat this procedure with the separated Congo red and phenol red spots . 

There are many protocols and different types of platforms available, e.g. GeneChips from Affymetrix, Illumina Bead Arrays, Arrays from Agilent, Applied Biosystems, GE Healthcare, customized spotted cDNA microarrays etc. the basic procedure for a large-scale measurement of gene expression involves the preparation of total or mRNA from the biological sample(s) under investigation (e.g. candidate tissue) and the hybridization of copied labelled RNA or cDNA to the DNA elements on the array surface (Fig. 1). 

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