What would happen if the anti-human Ig-conjugate is not washed free of the well before the substrate is added?

Question:What would happen if the anti-human Ig-conjugate is not washed free of the well before the substrate is added?

1.The ELISA would not develop when the substrate was added

2.The ELISA would develop normally

3.All wells would show uniform over-development due to unbound and excess anti-human Ig enzyme conjugate

4.None of the above


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