ELISA
It allows one to determine the composition of cells with the naked eye. It turns antibodies, proteins of the immune system that identify and help attack pathogens, into detection tools. It’s tedious, it’s repetitive, but its payoff is tremendous. It’s the science behind the pregnancy test, HIV test, and more beyond imagination.
This is ELISA, an Enzyme-Linked Immunosorbent Assay.
We did an ELISA with a patient sample who had breast cancer to determine its type. The types are distinguished by the type of receptors located on the cancerous cells. There are three types of receptors: estrogen, progesterone, and HEL2. The type of breast cancer would point to its possible treatment. If the patient was estrogen and progesterone receptor positive, then hormone therapy would be in order. If not, hormone therapy wouldn’t do anything, and would only cause additional strain on the body.
To determine the patient’s type of breast cancer, we used antibodies. Antibodies bind to an antigen, a region of a pathogen. To detect if a certain antigen is present in a sample, one can add specifically-designed antibodies and see if they bind to them. However, this is happening at the sub-cellular level. How do we know if the binding has occurred?
We added a second antibody to bind to the first one, as seen in the picture on the
left. This secondary antibody was bound to an enzyme, which is a protein that
catalyzes a reaction with a substrate. We, thus, then added a substrate. The enzyme
formed a complex with the substrate, and their reaction produced a color change. Thus, as we
added the substrate, those with the antigen turned blue, while those without them stayed clear. This is how we were able to see which receptors the patient sample had, as seen in the picture on the right.
When the receptors present on the cancerous cell bound to their respective proteins, the cell would receive the signal to divide. This continuous division eventually forma a tumor, which could potentially spread throughout the body. The relationship between these surface receptors and the uncontrollable cell division characteristic of cancer cells shined more light on the nature of cancer for me.
Another aspect of this lab that I appreciated was what it demonstrated about experiment protocol. Whenever one runs an experiment, one needs a treatment and a control group. The control does not receive the treatment, so one can accurately compare their results. In this case, however, there were two controls: a positive and a negative one. The positive one contained all three receptors, while the negative one contained none of them. In this way, one could compare the patient sample to both, to ensure one wasn’t misinterpreting the data. In addition, everything was done three times over to ensure there wasn’t a fluke. All of this broadened my understanding of experimental design.
Truly, this experiment was exciting; everything was happening at the microscopic level, so we didn’t know if it was going to work until the final step. The ELISA experiment was definitely the highlight of my time at the Pathways Research Explorers program at the Fred Hutch.
Comments