Measuring Stomatal Density in Rice

Materials and Reagents

1. Microscope cover glasses (Matsunami Glass, 24 x 40 mm No.1)
   Note: Though tougher glass slides (1-1.2 mm thickness) are safer, you should use cover glasses if the objective lens of the microscope is designed for standard 0.17 mm glass thickness. Only special objective lenses that have long working distances (ELWD, LWD) will allow observation through a thick glass slide. Alternatively objectives designed for use without a cover glass (NCG, NC) will enable direct observation of imprints placed on the slide glasses. I use large (24 x 40 mm) glasses for making imprints covering large leaf area. Larger glass is also useful when you set it on the microscope stage.
2. Instant glue (Aron Alpha Super Set) (Toagosei Co., catalog number: EA936A-5 )
   Note: Aron Alpha Super Set includes liquid glue and accelerator. Aron Alpha is sold as Krazy Glue in North America and Cyanolit in Europe.
   
3. Accelerator for instant glue
   Note: While I use the genuine accelerator made by Toagosei, third-party accelerators developed for Aron Alpha (Krazy Glue) will work fine. You can observe stomata without the accelerator, but the imprints may be less clear.
   

Equipment

1. Light microscope with equipment for photomicrography
   
2. Eyepiece micrometer and stage micrometer
   Note: These are indispensable for measurement of field of view unless your microscope can automatically calculate scale bars.
   

Procedure