Page 353 - The Veterinary Laboratory and Field Manual 3rd Edition

322 Susan C. Cork, M. Faizal Abdul Careem and M. Sarjoon Abdul-Cader

that requires accurate and precise readings an • For high throughput screening, some sys-
ELISA plate reader, which determines the spe- tems are not sufficiently robust to give good
cific absorbance level for the colour change in repeatable results under less than perfect
the test, should be used. The ELISA plate reader conditions (for example, water quality may
can be calibrated to give an assessment of ‘titres’ have to be high and consistent).
for test samples by using titrations of known • Local conditions may affect results (voltage
positive samples along with negative controls fluctuations, low voltage, power cuts, poor
and preparing a standard curve or assessing refrigeration capacity and unreliable storage
values against a predetermined ‘cut off’ point. of reagents, availability of a generator).
The plate reader will record optical density read- • Ambient temperatures inside working labora-
ings and calculate which samples have readings tory rooms may fluctuate and can be variable.
above the ‘cut off’ point, that is, those that are This may be a factor to consider.
likely to be positive. In single sample test kits the • Tests done in small numbers will create waste
result is usually clear cut with a colour change (except where single sample kits are used).
observed in the positive control and a similar • Kits do not always contain all the required
reaction seen for positive samples. Negative items and replacement consumables and
samples will usually show no colour change or reagents may not be available locally.
a different colour change to that of the positive • Many kits have not been fully validated
control. The instructions provided with the kit for use on field samples from animals of
tests will outline what to expect. unknown disease status, that is, cut off levels
ELISA remains one of the most sensitive and and potential cross reactivity with antigen–
specific of the serological tests carried out at antibody used in the kits. Kit performance
the level of regional laboratories although the may need to be further examined by under-
relative degree of sensitivity or specificity var- taking base line screening on samples from
ies with the test used. In this context, the term the population(s) of interest and comparing
‘sensitivity’ is the ability of the test to detect the ELISA result with those of a gold stan-
true positive samples and the term ‘specificity’ is dard test, that is, AGD, CFT, HI and so on.
the ability of the test to distinguish between true
positive and negative samples. In some tests, the Nonetheless, ELISA test technology is a huge
sensitivity is increased at the expense of specific- step forward in the diagnostic capacity of labo-
ity, that is, all positive samples are detected but ratories in developing countries.
some false positives also occur. These tests are ELISA test kits and systems now come in a
useful when the aim is to detect and eliminate a variety of formats but essentially; in the ELISA
disease, but in the later stages of a disease con- enzymes are coupled to antibodies to mark or
trol programme it is often necessary to re-test highlight immune complexes. The subsequent
suspected ‘false positive’ cases using other more action of a substrate specific for the enzyme
specific methods of diagnosis. The ELISA is will then produce a colour reaction indicat-
becoming one of the most widely used serologi- ing the presence and strength of the immune
cal tests for rapid diagnosis of animal diseases complex.
especially in surveillance programmes, but the A solid phase system is usually used for the
following disadvantages must be noted. test which involves binding protein (antigen or
antibody) onto a polyvinyl or polystyrene surface
• The cost of the kits can be high and purchase as a first step. This can take place in the well of
may require foreign currency. a microtitre (96 well) plate (8 × 12 well strips)

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