Longitudinal cross section of the gynoecium from a May Apple ( Podophyllum sp.) showing a single ovule and a developing embryo. The Safranin-O in the Flemmings triple stain has stained nuclei red. The orange G has stained cell walls with much greater differentiation than obtained from Hematoxylin-Erythrosin-B. The gynoecium as collected is at www.flickr.com/photos/14643312@N02/18069047169. Specimen from the edge of the Dowagiac River (5/15).
The following protocol is largely based on Chamberlain, "Methods in Plant Histology, 5th ed., 1905. Specimens fixed in FAA (formaldehyde, acetic acid, ethanol) 24 hr. Dehydrated in IPA at 35, 50, 70, 85, 91, 95, 99 %. Infiltrated with paraplast saturated xylene 48 hr followed by 2 Paraplast baths prior to embedding in Paraplast. Sectioned 11 um thick on a Spencer 820 microtome. Cleared in xylene, 5 min, 2X. Rehydrated 99, 95 % IPA,10 min. each. Stained with Flemming's Safranin, Crystal Violet, orange G. as follows. Stained 1% Safranin in 50% IPA 24 hr. Differentiated 50 % IPA, 10 min. Stained 1 % aq Crystal Violet 10 min. Dip 4X in water. Stain 1 % Orange B 1 min. Dip 4X 95 % IPA, agitate 5 sec 99 % IPA. Rinse off IPA with Eugenol, wick off. Pour Eugenol over slide. Cleared 2X xylene 2.5 min each. Mounted with DEPEX.
Photographed on a Spencer 42 petrographic polarizing microscope using an original magnification of 430X, using a Sony NEX 5N with a Leica MIKAS 1/3X adapter.
Longitudinal cross section of the gynoecium from a May Apple ( Podophyllum sp.) showing a single ovule and a developing embryo. The Safranin-O in the Flemmings triple stain has stained nuclei red. The orange G has stained cell walls with much greater differentiation than obtained from Hematoxylin-Erythrosin-B. The gynoecium as collected is at www.flickr.com/photos/14643312@N02/18069047169. Specimen from the edge of the Dowagiac River (5/15).
The following protocol is largely based on Chamberlain, "Methods in Plant Histology, 5th ed., 1905. Specimens fixed in FAA (formaldehyde, acetic acid, ethanol) 24 hr. Dehydrated in IPA at 35, 50, 70, 85, 91, 95, 99 %. Infiltrated with paraplast saturated xylene 48 hr followed by 2 Paraplast baths prior to embedding in Paraplast. Sectioned 11 um thick on a Spencer 820 microtome. Cleared in xylene, 5 min, 2X. Rehydrated 99, 95 % IPA,10 min. each. Stained with Flemming's Safranin, Crystal Violet, orange G. as follows. Stained 1% Safranin in 50% IPA 24 hr. Differentiated 50 % IPA, 10 min. Stained 1 % aq Crystal Violet 10 min. Dip 4X in water. Stain 1 % Orange B 1 min. Dip 4X 95 % IPA, agitate 5 sec 99 % IPA. Rinse off IPA with Eugenol, wick off. Pour Eugenol over slide. Cleared 2X xylene 2.5 min each. Mounted with DEPEX.
Photographed on a Spencer 42 petrographic polarizing microscope using an original magnification of 430X, using a Sony NEX 5N with a Leica MIKAS 1/3X adapter.