This device consists of a siphon system and 5 or 10 grid disks, modified from the previous mode , for large numbers of grids with ultrathin sections. This method improves the ease of assembling grids onto the grid disk and also requires much less stain solution. This system only takes 5 min for one single stain washing, at a maximum of 100 grids, and also avoids stain contamination. The grid disk can also be used for immunocytochemistry work and for critical point drying of grids with biological specimens.
MATERIALS & METHODS
This autowasher set shown in Figs.
A & B is made up of a glass washing chamber, grid disks, and a disk holder. The glass chamber (15 mm I.D. x 60 mm Long) with a siphon tube (5 mm I.D. x 50 mm Long) allows stain solution to be retained up to 6 ml which can then be quickly replaced by distilled water during washing process. This system allows grids to be washed continuously when the water flows at an appropriate rate. The disk holder consists of a glass tube (4 mm O.D. x 100 mm Long) with its top end attached to a rubber stopper, two small disk stoppers, and one grid guard. On the rubber stopper a venting air opening is made to allow free flow of water. The disk holder holds up to 10 grid disks. Each plastic grid disk (4 mm I.D. x 4 mm High, Fig. ) has 10 open slits to hold 10 grids. Thus, using this system, a total of 100 grids can be washed simultaneously.
Staining and washing:
- To load grids on a disk: Insert the wider glass tube (5 mm O.D. x 90 mm Long) into the disk as shown in Fig . This will open the slits to enable grid insertion. Follow the triangle-shaped cutting edge on the disk to insert a grid into the open slit (Fig. ). After loading all 10 grids remove the glass tube with forceps while holding the disk down (Fig. ).
To assemble grid disks to the disk holder: Each disk holder can hold 10 disks. To assemble the disks, simply remove the bottom stopper and grid guard from the holder (Fig. ). Then insert the disks in order according to the alphabetic labeling. Finally, put the grid guard and the stopper back on the holder.
3. Staining:
The amount of stain solution will depend on the number of disks in the holder. Ideally, the glass chamber should be filled with enough stain solution to just submerge the disks in the washing chamber. In general, 2 ml is required for the first disk and 1 ml is required for each additional disk. Then attach the washing chamber to the disk holder. This device can then be attached to the distilled water tank (Fig. ).