Superfrost* plus slides

Any archived formalin-fixed, paraffin-embedded tissue blocks can be used for preparing tissue sections. However, there are cases that would never demonstrate successful BISH (or FISH) signals. Since, in general, information of how tissue samples were processed for paraffin embedding is very difficult to obtain, the exact cause of unsuccessful BISH assays cannot be clearly identified. The delay of placing tissue samples in a fixative is known to create difficulties of successful molecular histopathology assays. Tissue sections should be cut at 4-5 Xm and placed onto SuperFrost Plus slides (Erie Scientific Company, Portsmouth, NH, USA). The tissue sections can be stored in a slide box at room temperature. 

Mount sections on microscope slides for processing with antibodies. Cryostat sections can be collected on microscope slides SUPERFROST PLUS slides hold sections well. Alternatively, glass microscope slides can be coated with a 0.1 mg/ml poly-L-lysine (Sigma PI399) solution in water. Loss of sections during processing is a frequent problem. 

OCT (TissueTek) and gelatin-subbed slides or Superfrost Plus slides (Fisher). 

Cut ribbons of 6-pm sections on a microtome. Float the ribbons of sections on a bath of distilled water at 40°C until free from creases. Collect on Superfrost plus slides. Dry slides at 37°C overnight. 

Cut section on a cryostat onto Superfrost plus slides... 

Mount the slices on Superfrost Plus slides, cover with Fluoromount , and seal slides with nail polish. 

Glass microslides should be precoated to facihtate adhesion. Use 0.1% poly-L-lysine (w/v) in distilled water, or purchase Superfrost Plus Treated Microscope Slides from Fisher Scientific (Pittsburgh, PA). Either works equally well. 

Excessive apphcation of tissue adhesive, gel-coated slides, or the use of albumin when preparing cytospins may contribute to background stain. The use of charged slides (Fisher Superfrost/Plus, Fisher Scientific, Pittsburgh, PA) can eliminate this problem. 

Kidney tissue is fixed with paraformaldehyde-lysine-periodate by vascular perfusion (Brown et al., 1996). Tissue slices are further fixed overnight at4°C with the same fixative and stored in PBS (pH 7.4) containing 0.02% sodium azide. They are placed in 30% sucrose in PBS for at least 1 hr, and then surrounded by a drop of Tissue-Tek embedding medium on a cryostat chuck before freezing by immersion in liquid nitrogen. Cryostat sections about 5 p,m thick are cut at a chamber temperature of -25°C, collected on Fisher Superfrost Plus charged slides, and stored at —20°C until use. 

Tissues are fixed in formalin, embedded in paraffin, and sections (5 xm thick) are transferred onto Superfrost Plus-coated slides (Mason et al 2000). The sections are deparaffinized with xylene and then rehydrated with descending concentrations of ethanol. They are placed in 0.1 M sodium citrate buffer (pH 6.0) and heated in a microwave oven at full... 

Specimens from biopsies, excisions or resections must be handled as soon as possible to preserve the tissue for FISH. Specimens should be preserved in 10% neutral-buffered formalin (NBF), preferably as 3-4 mm blocks fixed for 18-24 hours followed by dehydration and embedment in paraffin. Sections should be cut into 4-6 pm, mounted on positively charged slides (e.g. SuperFrost Plus, Mentel-Glaser, Thermo Scientific) and adhered to the slide by baking at 60°C for approximately 1 hour. 

Tissue sections of 5 pm thickness. Paraffin sections mounted on commercially available silane-coated glass slides, such as standard SuperFrost Plus (Fischer Scientific, Pittsburgh, PA) or on slides prepared using a home-made APES coating or frozen sections mounted on silanized, gelatin-coated or poly-L-lysine-coated slides (see Notes 1-4). 

For optical imaging, a fifth section is cut from embryo 3 and is designated as section 11. This section is captured with a clean SuperFrost/Plus glass slide. 

Glass slides. For some tissues, it is helpful to first treat slides with agents that improve adhesion, including polylysine solutions, aminoalkylsilane, and gelatin "subbing" solutions. Commercially available treated slides such as Superfrost Plus (Fisher) may also be useful. 

Superfrost Plus Microscope Slides (Thermo Fisher Scientific Inc., Waltham, MA). 

Adhesion to slides can be improved by using commercially available slides with increased adhesiveness, e.g., Superfrost Plus and ColorFrost Plus Microscope Slides, Thermo Scientific Inc., Waltham, MA. 

J,m should be mounted onto 3-amino-propyltriethoxysilane-coated (APES Sigma, St. Louis, MO) or charged (Plus Superfrost slides Fisher Scientific, Pittsburgh, PA) glass slides and attached by overnight drying at 58°C. 

Alternatively, charged glass slides, such as SuperFrost (Ultra) Plus (e.g. Fisher Scientific), provide highly standard adhesion even under extreme HIER conditions. Heat activation of the binding between the adhesive layer and the tissue section by melting the wax, for at least 30 min, is crucial before dewaxing. 

Tissue sections mounted on adhesive glass slides, e.g. SuperFrost (Ultra) Plus (Fisher Scientific) and heat activated at 60°C (3-5-p.m thick paraffin sections) or at 85°C (l-2- xm thick epoxy resin sections) for at least 30 min. 

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