In neuroscience and other biomedical research fields, animal models allow us to answer critical but otherwise impossible questions. Despite the value of these models, however, sometimes nothing can replace the real deal—human tissue.
The limitations of human tissue research stem from the variability between subjects and the risk for covariates that influence your results. Which are all true—humans are messy. But let’s be honest, how boring would life be if we were all the same? This doesn’t make human tissue research impractical. We just have a lot more to consider when performing experiments. If you properly address all of these potentially confounding factors, though, human tissue studies can provide invaluable information.
Below are things to know and consider when working with human tissue.
Humans Are Messy
Consider the differences between the individuals in your study from the experimental to the interpretation of results. If you just include only the most basic variables (age, race, sex, postmortem interval, and tissue pH), you leave out a lot of important information. However, the list of potential variables goes on forever—narrow them down to the most important factors that affect your data. In disease studies, for example, the age of onset, duration, and medication history might be important to include. If you don’t know where to start, look up previous human studies in your field to see the variables other researchers included.
When comparing multiple cohorts, match your experimental and comparison subjects on the most essential factors. Then, include the variables in your statistical analyses to check their influence on your results.
Preservation of Human Tissue
Just because you saw it in a creepy horror movie, doesn’t mean the IRB (Institutional Review Board) will find it appropriate to perfuse human subjects. Animal tissue has the advantage here. It can be frozen in time without the damaging effects of a postmortem interval.
Human tissue is typically preserved by freezing it or immersing it in a fixative solution, such as 4% paraformaldehyde. If you receive tissue at an early stage, experiment to find an ideal fixation time. You want one that gives great preservation but does not overfix the tissue and mask the antigen you want to study.
In addition, you can perform initial dissections to reduce the thickness of the tissue. Decreasing the tissue thickness will preserve the tissue faster and better maintain structural integrity.
Transitioning from animals to humans requires you to scale up your experiments. The “small” brain region I studied in human tissue dwarfed an entire hemisphere of a rat brain. Thus, it may no longer be possible to perform immunohistochemistry on your entire set of sections at once. Instead, perform your experiments in batches to produce high quality staining in each section. Process your comparison groups together (an equal number from each group per batch) to minimize methodological differences between groups.
I already touched on the downside of human tissue preservation for structural integrity, but it’s important to consider this for protein studies as well. Immediately after death, proteolysis gets busy breaking down proteins in soft tissues. Depending on the protein and postmortem interval length, the perimortem protein concentration could decrease over half by the time the tissue is preserved. Therefore, the total protein concentration that gives you beautiful western blot bands with rat tissue will likely not be adequate to give a strong signal in human tissue. Optimize your protein study to use the least amount of tissue necessary but still achieve a nice signal.
When quantifying protein, know how the postmortem interval might affect your data. You might be lucky enough to find a study examining the length of the postmortem interval on your protein of interest. Even so, be sure to include the postmortem interval as a covariate during your analysis.
Human tissue is critical to biomedical research. Knowing how to best use the tissue and properly address the concerns unique to human studies can help you get the most out of each valuable donation and produce solid human tissue research.