Hi Abhishek,

I am not a specialist for the cell types you are using - however I think both cell lines should only produce relative low amounts of IL-1ß ( and I assume also TNF alpha) when they are not activated e.g. due to LPS.

Regarding your question: Do you have a positive control in your realtime PCR which ensures for example that you have a good quality template?

Even if you put in the same amount of cDNA (how do you quantify this?) you can still have quality issues with your cDNA from the HMC3 cells.

Check if a positive control (you can use a house keeping gene like GAPDH or ß-actin) also have low CT values (or is not detectable).

If you have problems with detecting the positive control this would indicate a low quality cDNA (maybe due to a low quality RNA-template in the first place). If you have this problem you should prepare new template and new cDNA.

If you get good results for your postive control from the cDNA from your HMC3 cells the differences you see might be due to differences in the expression of your target genes between you cells lines and how you will go on with this should strongly depend on what you want to show in your experiment (what I don't know and I can't advice you on this).

However, I would not recommend to simple increase your cDNA amount without checking what might be the reason for the differences you see (e.g. bad quuality cDNA or differences in gene expression)

Good luck