Incubation preferred

Wipe excess liquid from the area around the sections, and apply sufficient 5% heat-inactivated normal serum from the second antibody species to cover each entire section (15 min). If protein A-gold probes are used, flood the sections with BSA-PBS rather than normal serum. It is important to keep the sections covered and in a humid environment throughout all incubations, preferably using a purpose-made humidity chamber. Depending on the size of the section, about 50-200 uL of reagent is sufficient to cover each preparation... 

Briefly stated, the production of chloramphenicol by the surface culture method involves inoculating a shallow layer, usually less than about 2 cm, of a sterile, aqueous nutrient medium with Streptomyces ver)ezuelae and incubating the mixture under aerobic conditions at a temperature between about 20° and 40°C, preferably at room temperature (about 25°C), for a period of about 10 to 15 days. The mycelium is then removed from the liquid and the culture liquid is then treated by methods described for Isolating therefrom tne desired chloramphenicol. 

A preferred method for the detection of pyrogens is the limulus amebocyte lysate (LAL) test. A test sample is incubated with amebocyte lysate from the blood of the horseshoe crab, Limulus polyphemus. A pyrogenic substance will cause a gel to form. This is a result of the... 

Incubate sections for 30 min with (strept)avidin conjugated with a fluorophore or with any enzyme (peroxidase or alkaline phosphatase). For localization of low-density antigens (<10K molecules/cell), VECTASTAIN ABC reagent (http //www.vectorlabs.com/) is preferable. 

As the enzyme itself is usually the focus of interest, information on the behavior of that enzyme can be obtained by incubating the enzyme with a suitable substrate under appropriate conditions. A suitable substrate in this context is one which can be quantified by an available detection system (often absorbance or fluorescence spectroscopy, radiometry or electrochemistry), or one which yields a product that is similarly detectable. In addition, if separation of substrate from product is necessary before quantification (for example, in radioisotopic assays), this should be readily achievable. It is preferable, although not always possible, to measure the appearance of product, rather than the disappearance of substrate, because a zero baseline is theoretically possible in the former case, improving sensitivity and resolution. Even if a product (or substrate) is not directly amenable to an available detection method, it maybe possible to derivatize the product with a chemical species to form a detectable adduct, or to subject a product to a second enzymatic step (known as a coupled assay, discussed further later) to yield a detectable product. But, regardless of whether substrate, product, or an adduct of either is measured, the parameter we are interested in determining is the initial rate of change of concentration, which is determined from the initial slope of a concentration versus time plot. 

For ligand-gated ion channel receptor expression studies, stage V and VI oocytes are preferred (see earlier). Under a dissecting microscope, oocytes of this stage that are in good condition are sorted from the rest and then incubated at 16-18°C in MBM overnight. Incubation in hypertonic phosphate... 

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