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Home / Techniques / Protocols / Toluidine blue (plants)

Techniques. Protocols

TOLUIDINE BLUE
(Plants)

Toluidine blue staining can be use in plant histology to get a quick staining. It is easy to prepare, and the procedure is very short. In addition, toluidine blue staining is very useful because it shows metachromasia, so that one dye stain with different colors depending on the tissular structure. The colors go from brilliant green to purple, through a range of blue.

The general staining for plant tissues is used to check the quality of tissues and structures. Although toluidine blue staining contains only one dye, others combined two or more, such as the classic safranine / Alcian blue.

Procedure

8 µm thick sections from fixed and paraffin embedded samples will be stained. The sections are attached to gelatin coated slides.

1.- 2 x 10 min in xylene

2.- 2 x 10 min in 100º ethanol

3.- 10 min in 96º ethanol

4.- 10 min in 80º ethanol

5.- 10 min in 50º ethanol

6.- 5 min in distilled H2O

7.- 30 s in toluidine blue (C.I. 52040) 0,5 % in cdestilled H2O.

8.- 3x10 s en H2O destilada con agitación

9.- 10 s in 96º ethanol, gentle agitation

10.- 2 x 10 s in 100º ethanol, gentle agitation

11.- 2 min + 5 min in xylene

12.- Mounted with mounting media and coverslipped

Results

Primary cell walls: blue to purple.

Secondary cell wall: brilliant green.

Cuticle: brilliant green.

Notes

This staining is quick and easy to do, particularly after the staining step. More or less staining intensity is obtained during the dehydration steps. Thus, it is convenient to dehydrate slide by slide and a gentle agitation to remove the dye homogeneously from the tissues.

Products

Xylene

50º, 70º, 80º, 90º, 96º and 100º ethanol

Toluidine blue (C.I. 52040), 0.5 % in distilled H2O

Distilled H2O

Mounting medium

Labware

Staining dishes

Slides racks

Coverslips

Toluidine ble
Section of a spine of common gorse (Ulex europaeus). The cuticle is stained with brilliant green, the sclerenchyma fibers with pale blue, and parenchyma cell walls in purple-violet.
Toluidine blue Safranin - Blue alcian
Common gorse (Ulex europaeus) stem. Toluidine blue on the left and safranin - Alcian blue on the right
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