ELISA (kit) Human Serum Cytokine measurements (Multiplex assay) -NA- Human

I used a serum/plasma kit for my serum samples. After the phase separation the samples should have 3 phases: a colourless aqueous phase, a white interphase and a red organic phase. However, in some of my samples there was no aqueous phase unless I wait for an extended period of time. How can I circumvent this problem?

Get tips on using Human Prolactin DuoSet ELISA to perform ELISA Human - PRL

Get tips on using Human MICB DuoSet ELISA to perform ELISA Human - MICB

Get tips on using Human Leptin DuoSet ELISA to perform ELISA Human - Leptin

Get tips on using Human LBP DuoSet ELISA to perform ELISA Human - LBP

Get tips on using Human GDNF DuoSet ELISA to perform ELISA Human - GDNF

Get tips on using Human Fibronectin DuoSet ELISA to perform ELISA Human - Fibronectin

Get tips on using Human Decorin DuoSet ELISA to perform ELISA Human - Decorin

Get tips on using Human Dkk-1 ELISA to perform ELISA Human - Dkk-1

ELISA is the most commonly used method of detecting and quantifying the concentration of an antigen in an unknown sample. During the experiment, If you get a weak signal, then make sure reagents are at room temperature before starting the assay. Try increasing incubation times to ensure maximal antibody binding and amplify the signal. Secondly, if you get values above 0 in the negative control indicates a high background signal. Try to consider reducing your antibody concentration and prevent non-specific binding of antibodies by using affinity-purified antibody and suitable blocking buffers. To avoid high well to well variation, do not stack plates during incubation, no bubbles in the plate and wash wells thoroughly to avoid variation.