Incubation chamber

Potentially mineralizable C and N are often measured by incubating a sample of field-moist soil at a known temperature in a sealed chamber containing an alkali trap. The C02-C accumulated in the trap is measured by acid titration and this represents the quantity of C mineralized. Alternatively, C02 in the headspace of the incubation chamber can be measured using a C02 analyser. The amount of N mineralized during incubation is calculated as the difference in extractable NH4+ - andNCV-N measured in the soil before and after incubation. Mineralizable N can also be measured in an open incubation system where the soil is leached periodically and NH4+- andNCV-N in leachates is measured (Stanford 1982). 

Self-contained incubation chambers are commercially available which can be used as either negative or positive pressure units. 

To assess evaporative loss of fluid from filled capillary gaps, clean slide pairs were filled with colored, distilled water. After a 1-min fill, the maximum height of fluid in the gap was marked on each slide pair. Slides were then segregated into two groups one remained exposed to room air, and the other was placed into the closed incubation chamber. At timed intervals, fluid loss in millimeters was recorded. Laboratory temperature was a constant 25°C with a relative humidity of 28%. After 40 min, air-exposed slides had an average loss of 33% of their volume, whereas slides in the incubation chamber lost only 9% in the same time interval. 

Because the 150- iL aliquots of antisera that are placed in the isolons are exposed to the room environment and suffer a potential 50% evaporative loss over 2 h, the optional hood should be required as standard equipment, a beaker of water should be placed in the oven, heated, and allowed to act as a humidity source, slide staining runs should be kept as short as possible, and the three-well reagent isolator should be used whenever possible because of its larger fluid capacity. To prevent excessive evaporative loss of fluids from the filled capillary gaps, it is imperative that the slides be placed in the incubation chamber having a proper seal. If left exposed to room environmental conditions, fluid evaporates from the capillary gaps with resultant reagent concentration. 

Sections (7 xm thick) of freshly frozen tissues are mounted on silane-coated slides and fixed with 4% buffered formaldehyde (pH 7.0) for 20 sec (Richter et al., 1999). The sections are rinsed in TBS (pH 7.4) for 15 sec, followed by incubation with EPOS antibody for 3 min at 37°C in an incubation chamber. They are rinsed twice for 15 sec each in TBS, and then developed with peroxidase-DAB detection kit (Dako) in a microwave oven (500 W) for 1 min during microwaving, the slides are cooled by a cold water bath (Werner et al., 1991). After being rinsed in tap water, the sections are counterstained with hematoxylin for lOsec. They are rinsed in tap water and cover-slipped. 

Remove the plate from the incubation chamber and place on ice, allow cooling for 15 min. 

After the operator has selected the desired method menu of the relevant samples and has started the instrument, all subsequent steps are fully automated. Since 1987 it is also possible to effect a direct identification of the sample so that there are no longer any problems in respect of a dialogue with a central EDP system. The samples are taken from the sample vessel by means of disposable single use pipette tips that are used for one sample only and exchanged via a computer-monitored pipetting unit. This method excludes the possibility of a carry-over between samples. In accordance with the preset conditions, the required slides are automatically moved to the sample dosage unit (see Fig. 23). Samples of 11 pi serum or plasma will be sufficient for kinetic measurements (enzymes), 10 pi of sample for all other tests. As soon as application of the sample has been completed, the slide is moved to the appropriate incubation chamber by means of the slide rotor (see Fig. 23). The chemical reactions take place in these chambers. This is followed by measurement either by reflectometer (end point or kinetic) or a potentiometric measurement unit. 

Emission rate in laboratory incubation chambers Calculated emission rate based on a defined area Calculated indoor air concentrations by a mold-infested site of 0.25 m ... 

Today, the company Affymetrix offers microarrays with >2,000,000 unique compounds. The fluidic system is quite simple. The sample is manually loaded with a pipette into the chip, and capillary forces transfer the sample to the incubation chamber. Incubation and mixing is enhanced by a moving air bubble actuated by slow rotation. 

Specific equipment microplate reader, incubation chamber (37°C). 

Remove the plate/s from the protective packaging and save the plastic holder for later use in the incubation chamber. 

The first category (Table 9.2), (1) Sample, collects the conditions of sample preparation. In the example here, kidney from a rat will be used following trans-cardiac perfusion. The tissue blocks will be infiltrated with 20% sucrose overnight, frozen in isopentane, and sectioned on a cryostat. For processing, the sections will be placed on a microscope slide. To calculate how much incubation and rinse solutions are needed, the size of the incubation chamber is listed. In the example, 250 p,l will just cover the tissue and can be used for each step. 

Incubation chambers/size Microscope slide 100 ml per area ... 

Incubation chambers/size 12 mm coverslip in 24-well plate ... 

Transfer 10-20 pg of protein from the Slide 1 Mix to the Slide 1 incubation chamber. Transfer an equal quantity of protein from the Slide 2 Mix to the Slide 2 incubation chamber (see Note 14). 

Prepare 50 ml Incubation Buffer from 45 ml Stock Incubation Buffer and 5 ml Background Reducer. Then, add 5 ml of Incubation Buffer to the incubation chambers. 

Transfer 200 pg of protein from Subheading 3.4 to each of the incubation chambers and incubate the tray at room temperature for 30 min with gentle rocking. 

Carefully remove the slides from the Storage Vial and place each one, label side facing up, into one of the two designated incubation chambers containing the native antigen mix. 

Remove the buffer from the incubation chambers and discard. 

Add 5 ml of PBS (pH 7.4) to the incubation chambers, taking care to pipette the liquid on the portion of the slide containing the manufacturer s label. Incubate at room temperature for 5 min with gentle rocking. 

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