Day 32: experiment juxtaposition
As I was talking about earlier this week, I've often been running more than one experiment at the same time during a week. But today, the differences between the two experiments seemed quite striking!
The two experiments are to look for two different indicators to suggest that my protein of interest is involved in making my cell type increase during cell stress/inflammation:
1. Look for the protein itself.
This involves separating the proteins using electrophoresis (pulling the proteins through a polyacrylamide gel using an electric current to draw the proteins through the tiny pores of the bell - the smaller proteins get to the bottom of the gel quicker as they can wiggle through the holes faster. Then using a method called Wet Transfer, which pulls the protein out of the gel and onto a PVDF membrane - a technique that uses litres of buffer.
2. Looking for the genes that are expressed - suggesting that the cell is producing the proteins and using the part of the genome that controls the production/expression of my protein of interest.
A really delicate experiment - requiring very accurate pipetting, careful regulation of conditions and uses tiny amounts of cDNA, probes and buffers. The experiment is carried out in a special 96 well plate and put into a sensitive machine which can detect the changes in fluorescence made by the probes attached to the genes of interest.
Running the two together seemed to highlight their differences - both refined techniques, but one seemingly a rudimentary and practical science and the other a much more contemporary technique requiring more precision. Felt out of place to be hauling a transfer tank out of a sink full of ice and in the next minute carefully placing 5ul of cDNA into the wells of a PCR plate!
The two experiments are to look for two different indicators to suggest that my protein of interest is involved in making my cell type increase during cell stress/inflammation:
1. Look for the protein itself.
This involves separating the proteins using electrophoresis (pulling the proteins through a polyacrylamide gel using an electric current to draw the proteins through the tiny pores of the bell - the smaller proteins get to the bottom of the gel quicker as they can wiggle through the holes faster. Then using a method called Wet Transfer, which pulls the protein out of the gel and onto a PVDF membrane - a technique that uses litres of buffer.
2. Looking for the genes that are expressed - suggesting that the cell is producing the proteins and using the part of the genome that controls the production/expression of my protein of interest.
A really delicate experiment - requiring very accurate pipetting, careful regulation of conditions and uses tiny amounts of cDNA, probes and buffers. The experiment is carried out in a special 96 well plate and put into a sensitive machine which can detect the changes in fluorescence made by the probes attached to the genes of interest.
Running the two together seemed to highlight their differences - both refined techniques, but one seemingly a rudimentary and practical science and the other a much more contemporary technique requiring more precision. Felt out of place to be hauling a transfer tank out of a sink full of ice and in the next minute carefully placing 5ul of cDNA into the wells of a PCR plate!
Comments
Post a Comment