Can the rat IL-6 ELISA measure both serum and tissue homogenates simultaneously?

Sixty samples is not a small number. First, take 3–5 representative serum and liver homogenate samples for a preliminary experiment: test the neat sample and 2/5/10/20-fold dilutions to determine the optimal dilution factor, then perform spike recovery. In the formal experiment, measure serum and tissue samples on separate plates as much as possible, but include a standard curve and two quality controls on each plate. Tissue homogenate samples tend to have high CV in replicate wells; it is recommended to aliquot and freeze the homogenate after homogenization to avoid repeated freeze-thaw cycles. Before loading, bring samples to room temperature and centrifuge at low speed to remove precipitate. Inter-plate variation can be monitored by repeating one quality control sample across different plates within the same batch.

The cost per well can basically be kept within 15 RMB for domestic 96T kits. For rat IL-6, you can focus on asking Lianke, Dayou, Sizhengbai, and Xinbosheng, and have their technical support provide application data or references for tissue homogenate samples. Before purchasing, confirm whether the sample types listed in the instructions include "tissue homogenate/tissue lysate," and whether the standard curve range covers your expected values. If a brand has only been validated for serum/plasma, it is not recommended to directly test tissue unless you perform full parallelism and spike recovery yourself. Even if the budget is tight, do not choose unknown OEM brands, as the cost of troubleshooting problems will be higher.

It is acceptable to use the same kit for measuring serum and liver tissue homogenate, but do not use lysis buffer containing strong detergents for tissue homogenization. High concentrations of SDS and Triton X-100 can interfere with antigen-antibody binding, causing the standard curve and homogenate matrix to be non-parallel. It is recommended to use PBS plus protease inhibitors or a mild lysis buffer recommended in the kit instructions. After homogenization on ice, centrifuge at 4°C, 10000×g for 10 minutes and collect the supernatant. It can be filtered through a 0.22 μm membrane to remove lipids. Liver tissue homogenate IL-6 levels correlate with the degree of inflammation. First measure BCA total protein, and finally normalize the data to pg/mg protein, which is more comparable than using pg/mL alone.

The impact of detergents mainly depends on concentration. If a lysis buffer containing detergent has already been used, prioritize diluting 5-20 fold to reduce the detergent concentration, while diluting with the kit's diluent. The method to assess interference is to perform parallelism: dilute the homogenate supernatant in a 2, 5, 10, 20-fold series and check whether the measured concentration remains stable after dilution. If the calculated concentration drifts significantly with increasing dilution factor, matrix interference is present. Serum is generally used undiluted or at 2-fold, while tissue homogenate is recommended to start at least at 5-fold. In acute inflammation models, liver tissue IL-6 may be high, so do not measure directly undiluted.