You most likely have evaporation. Try centrifuging the tubes in case there is liquid in the underside of the cap.

Another possibility is that your micropipettes are not calibrated properly. But that seems unlikely if you are noticing that the reactions appear to be equal volumes and then become unequal.

I'd first try making sure the caps snap tightly and are sealed on each of your tubes. Then try another brand of PCR tubes. Finally, a really old-school (and kind of messy) solution is to put a small volume of mineral seal oil on top of each PCR reaction. Make sure the oil has a high flash point (above 100*C) to avoid fire danger.

Try to use another type of PCR strips. the one you are using might not bear high temperature.

One possibility is that the missing water is condensed on the cap of the tube.

Another is that your tube seals are not working and some liquid is being lost.

Another is that the hot air at the end of the pcr process is holding more water than the colder air when you were setting up the reaction but that would only be 1ul or less The most likely reason is leakage which can come from screwing down the heated lid too tightly and distorting the tube/strip/cap allowing vapour leakage during cycling

You most likely have evaporation. Try centrifuging the tubes in case there is liquid in the underside of the cap.

Another possibility is that your micropipettes are not calibrated properly. But that seems unlikely if you are noticing that the reactions appear to be equal volumes and then become unequal.

I'd first try making sure the caps snap tightly and are sealed on each of your tubes. Then try another brand of PCR tubes. Finally, a really old-school (and kind of messy) solution is to put a small volume of mineral seal oil on top of each PCR reaction. Make sure the oil has a high flash point (above 100*C) to avoid fire danger.