What type of spot detection systems are available?
Horseradish peroxidase (HRP), alkaline phosphatase (AP) or gold particles can be used as streptavidin conjugates. The advantage of using HRP is its fast turnover rate (spots develop fast), whereas the drawback is increased background. The HRP substrate AEC (3′-amino-9-ethylcarbazole) forms intense red colored spots. However AEC is unstable and spots will bleach in a short period of time. AP has a linear reaction rate with little risk of high background. However spot development is slow and bleaching of spots is a problem. A highly effective way of spot staining can be achieved with gold-labeled streptavidin using silver precipitation for spot visualization. Because of the high stability of silver, spots do not fade and ELISPOT plates can be reanalyzed after being stored for several years at room temperature.
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