Introduction

During preparation for a class in histology at Victoria University a triple stain was sought for the distinction of muscle and connective tissue that would give consistently good results using stains that are usually readily available in a department of zoology. The material to be stained was sections of vertebrate tissue, in particular that of mouse and chick embryo. A staining method giving green connective tissue, pinkish cytoplasm and muscle, and blue nuclei was wanted. Gurr (1962) gives the following stains for obtaining a very similar staining reaction, viz. Weigert's haematoxylin, eosin yellowish, and naphthol green B. His method is as follows:

1.	Paraffin sections are mounted on the slide and brought down to distilled water.
2.	Stain for 6 mins. in a freshly prepared mixture of equal volumes of Weigert's haematoxylin A and B.
3.	Wash thoroughly in tap water; then stain for 3 mins. in 1% aqueous eosin y.
4.	Wash in tap water; then immerse in a solution of 10% ferric chloride for 5 mins.
5.	Rinse well in distilled water; then stain for 5 mins. in a 1% aqueous solution of naphthol green B.
6.	Differentiate for 2-3 mins. in 1 % acetic acid.
7.	Drain well; then dehydrate with acetone, afterwards clearing in acetone-xylol; then mount.

With the above procedure results were far from consistent and a clear differentiation of muscle and connective tissue was not obtained.

Testing the staining procedure consisted of working through the schedule using the times given by Gurr, and varying the times in the eosin, ferric chloride, naphthol green B, and acetic acid baths. Material used was F.A.A. fixed paraffin embedded sections of (a) mouse head, abdomen and duodenum, cut at 8μ on a rotary microtome, and (b) sections of frog stomach cut at 6μ on the same microtome. Staining procedures using alcoholic eosin as the plasma stain, and connective tissue stains of naphthol green B (George T. Gurr, 09506) in absolue alcohol and fast green FCF (Flatters and Garnett Ltd.) in absolute alcohol, clove oil, and cellosolve (ethylene glycol monoethyl ether) were also tried.