Ch. 8— Alternative to Animal Use in Testing • 183 
the reproducibility of the test itself, this might be 
satisfactory for some purposes, and it certainly 
warrants further investigation. Furthermore, many 
of the larger deviations in this study, upon fur- 
ther examination, were found to involve report- 
ing errors. This program relied on a multi-tiered 
classification scheme based on chemical structure 
(36). 
SKIN AND EYE IRRITATION 
The widely used Draize eye irritation test and, 
to a somewhat lesser extent, the skin irritation test 
have been criticized because of the amount of pain 
inflicted and because they are unsatisfactory mod- 
els for human irritation (91,95). First, the rabbit 
eye has structural differences, such as a thinner 
cornea apd differing tearing apparatus (103), and 
animal skin is much less sensitive and discriminat- 
ing than human skin (56,63). Second, both of these 
tests are sensitive to too many variables, making 
reproducibility poor (83,118). 
As with most tests, the number of animals used 
can sometimes be reduced. Several refinements 
have also been proposed. For example, screening 
tests based on pFI or skin irritancy might also serve 
as alternatives to eye irritancy tests in limited cir- 
cumstances, although preliminary studies indicate 
that this approach is frequently misleading (119). 
Other refinements involve local anesthetics (51,65, 
110), applying smaller (43) or more dilute (120) 
doses, and testing whole eyes in vitro (20). The lat- 
ter method has particular appeal when cow eyes 
are used because they are so readily available from 
slaughterhouses. In the case of smaller doses, a 
recent comparison with over 500 accidental human 
exposures showed that doses smaller than those 
now in use yielded results more predictive of the 
human response while causing less severe irrita- 
tion (38). 
Skin and eye irritation are similar in many re- 
spects . Thus, even though little work has been done 
to develop alternatives to skin irritation tests, the 
many approaches just summarized for eye irrita- 
tion may eventually be applied to skin testing as 
well (91). 
In Vitro Tests 
Several in vitro alternatives have been examined, 
and it appears to some commentators that no sin- 
gle alternative will be adequate, but that a battery 
of in vitro tests might be a useful replacement (67). 
Several types of cell cultures have been used in 
developing an in vitro test for eye irritation. The 
cells used are rabbit and human corneal cells (72), 
mouse and hamster fibroblasts, human hepatoma 
cells, and mouse macrophages (96). 
A variety of effects have been used as surrogates 
for eye irritation, such as the rate of uptake of uri- 
dine as an indication of cell functioning and re- 
covery, visible changes in cell structure, decreases 
in the concentration of cell protein (96), and re- 
lease of plasminogen activator from the injured 
cells (21). Some techniques appear promising, par- 
ticularly in their ability to rank substances based 
on irritancy. Rapid progress is being made in the 
development of techniques, but none can be con- 
sidered validated at this time (91). 
To date, little work has been done on in vitro 
replacements for skin irritancy testing. However, 
the growth of skin in tissue culture is of interest 
for treating burn victims, and it is expected that 
culture techniques currently being developed for 
that purpose can be used in testing methods. In 
addition, it has also been suggested that suitable 
specimens can be obtained from cadavers and 
surgery and from judicious use of human volun- 
teers (63). 
Chick Embryo 
One test system receiving considerable attention 
is the fertilized chicken egg. A part of the eggshell 
is removed and the test substance applied to the 
chorioallantoic membrane surrounding the devel- 
oping embryo (see fig. 8-1). This test has the po- 
tential for assessing both eye and skin irritancy. 
The chorioallantoic membrane of the chick em- 
bryo is a complete tissue, including arteries, capil- 
