214 
Psyche 
[September 
of the amount of variability between the patterns of individual 
spiders. 
The spiders used in this study were mature females of the sibling 
species Latrodectus mactans (Fabricius) and Latrodectus variolus 
Walckenaer collected in Florida during the summer at St. Petersburg 
and Torreya Ravine State Park respectively. These females are very 
difficult to separate using conventional morphological criteria. 
Hemolymph was obtained from individual spiders by severing the 
legs at the junction of the coxa and trochanter and then drawing up 
the expressed hemolymph into a polyethylene capillary tube connected 
to a i cc hypodermic syringe. The hemolymph was then mixed with 
O.05 ml of a Karo syrup — buffer solution (9 parts Tris buffer pH 
9.0: 1 part Karo syrup). The Karo increased the density of the 
solution making it easier to layer the sample into the gel shots. 
The electrophoretic runs were carried out in a cell manufactured 
by the E-C Apparatus Corp. in which a 8-slot, 8 per cent acrylamide 
gel slab was used. The hemolymph sample from each spider was 
loaded into one of 6 slots, the other 2 slots were loaded with 0.1 ml 
of a solution of human serum albumin (0.05 mg/ml). The albumin 
solution was lightly stained with bromthymol blue so that its migra- 
tion could be followed. Tris buffer (pH 9.0) was used in the buffer 
compartments of the cell and 150 ma (560-780V) was applied for 
90 min. The gel slab was then stained with amido schwarz and the 
relative mobility of each of the hemolymph protein fractions was 
obtained by comparing its mobility to that of the human serum 
albumin. 
Results 
The individual electrophoretic patterns of the hemolymph proteins 
of 10 L. mactans and 11 L. variolus females were recorded. Five 
fractions were found in each species and the average relative mobility 
of each of these fractions is given in Table I. Each fraction is num- 
bered relative to its position from the origin, fraction 1 being the 
closest to the origin. Figure 1 shows the characteristic patterns of the 
two species. All were run in the same gel. There are no significant 
differences in the average relative mobilities of fractions 1, 2, 4 and 
5 in the two species, but there is a clear cut difference in those of 
fraction 3. The magnitude of the difference is such that the relative 
mobility of this fraction can be used to separate all tested females of 
the two species. 
