Maule Staining for Lignin: A Technique to Characterize Lignin Distribution in Arabidopsis Thaliana Stem Sections

Abstract

Source: Pradhan Mitra, P. et al., Histochemical Staining of Arabidopsis thaliana Secondary Cell Wall Elements. J. Vis. Exp. (2014).

In this video, we describe the Maule staining technique for staining syringyl lignin present in the specialized plant cells. This staining technique helps to characterize the lignin distribution in the xylem and interfascicular fibers.

Protocol

1. Mäule Staining Dissolve 0.2 g of potassium permanganate in 40 ml distilled water to prepare a 0.5% potassium permanganate solution. Store in a dark bottle at room temperature for a maximum of 7 days. Prepare a 3.7% HCl solution by adding 1 ml concentrated HCl (37 N) to 9 ml distilled water (1:10). Prepare a fresh 3.7% HCl solution on the day of the experiment. Make sure that the 37 N HCl stock solution being used is not too old. Concentrated ammonium hydroxide solution (14.8 M) s…

Representative Results

Figure 1. Mäule staining of A. thaliana stem cross-sections. Mäule staining (red color) of a wild type A. thaliana stem section showing the normal lignin deposition in the walls of interfascicular fibers and xylem cells (A-E). The positions of the epidermis and cortex (Ep), interfascicular fibers (Fi), pith (Pi), and xylem (Xy) are indicated. Magnifications: Panel A: 5X; Panel B: 10X; Panel C: 20X; Panels D and E: 40X….

Materials

Agarose	EMD	MERC2125	CAS Number: 9012-36-6
Hydrochloric acid	EMD	HX0603-75	CAS Number: 7647-01-0
Vibratome	Leica	Leica Vibrating blade microtome VT1000S	http://www.leicabiosystems.com/ products/sectioning/vibratingblade-microtomes/details/product/ leica-vt1000-s/
Razor	American Safety razor company	Item # 60-0139-0000	Stainless Steel Double Edge Blade (Personna Super)
Screw Cap Microcentrifuge Tubes (2 ml)	VWR	16466-044
Microcentrifuge Tubes (0.6 ml)	Axygen Scientific	MCT-060-C
Tissue adhesive	Ted Pella Inc	10033	Store at 4 °C or 20 °C for 3 months or longer storage
Camera with CCD chip with no mechanical shutter	Hamamatsu	C4742-95
Camera software	Molecular Devices	MetaMorph version 7.7.0.0
Imagining anaylsis	Adobe	Photoshop CS4
Micro Cover Glasses, Square, No. 1	VWR	48366-067	22 x 22 mm (7/8 x 7/8")-Cover glasses are corrosion-resistant
Frosted Micro Slides, 1 mm	VWR	48312-003	75 x 25 mm – 1 mm
Ammonium hydroxide	EMD	AX1303-6	CAS Number: 1336-21-6
Potassium permanganate	Sigma	223468	CAS Number 7722-64-7