Optical Microscope Thickness Requirements for Glass Slides
Microscopic Dehydration and Sample Attachment: Due to repeated centrifugation, the cells are lost, so it is best to attach the cells to the surface of a glass slide or plastic sheet for optical microscopy, so that the cells are dehydrated and dried together with the glass slide or plastic sheet. Requirements for the thickness of glass slides in optical microscopy In order to allow cells to attach to glass slides or plastic sheets, it is necessary to first lay a layer of film on glass slides or plastic sheets. The method of laying the film is as follows:
(1) The simplest method for optical microscopy is to lay a thin layer of polyvinyl formal (formvar) film. Soak the cleaned slides in (0.5-1)% polyvinyl formal chloroform solution. After the atmosphere volatilizes, a white film appears on the slides. Shake the slides several times in the chloroform to make the film change. Thin and ready to use.
(2) Drop 0.1 latitude poly-L-lysine (dissolve 1 mg poly-lysine in 1 ml filtered water during preparation) on the cleaned glass slide, and put it in a 450C supply box after the droplet unfolds. dry. This method is best because poly-L-lysine is positively charged, allowing more surface negatively charged cells to attach.
(3) Add plasma and glycerol to distilled water at the ratio of (30--40)% and (3-5)% according to the requirements of the optical microscope for the thickness of the slide to form a glycerol protein solution. The liquid is dropped on the cleaned glass slide, unfolded and dried under the optical microscope. The laid protein film is immersed in 2% glutaraldehyde to fix the protein, washed and dried for later use. Before use, the plasma is dropped on the protein film Make it "activated", rinse with buffer after 30 minutes, and it can be applied directly.
The fixed and cleaned cells are made into a suitable concentration of cell suspension with filtered water under a digital microscope, and the optical microscope is placed on a glass slide or plastic sheet that has been covered with a membrane, and placed in a humid, low temperature (40C) environment ( 30-120) minutes to allow the cells to attach to the membrane. Use fine tweezers to pick up the glass slide and shake it in clean double distilled water, and wash away unattached cells with an optical microscope. Requirements for the thickness of slides in optical microscopes Polarizing microscopes are then quickly placed in a vessel filled with 50% acetone (or ethanol). The dehydration process is to put the slides into acetone or ethanol with increasing concentration every 3 minutes, and the optical microscope or continuously increase the concentration of the dehydrating agent in the glass slide container. According to the author's laboratory experience, the latter operation is more convenient. The specific method of optical microscopy can suck out half of the dehydrating agent in the container every 3 minutes, and then add an equal amount of dehydrating agent with a concentration of 100%. After 5-6 operations, the concentration of the dehydrating agent in the microscope price vessel reaches above 0.5%, and the cells are dehydrated successfully. After critical point drying, the glass slide or plastic sheet is glued to the sample pier with conductive adhesive for metal coating. There are also conveniences for attaching cells to membranes laid down by the third method. Optical microscope can hang the cells fixed by low concentration of glutaraldehyde on the "activated" protein film. The optical microscope requires the thickness of the glass slide to be placed for 10 minutes, and the glass slide or plastic sheet is soaked in 2% glutaraldehyde. After 30 minutes, rinse with filtered double distilled water, and then dehydrate, dry and metal coat according to the above method.
