Bio-friendly alternative to Xylene in histological staining procedures
General description of the project
My research project is centralized on testing pine oil as a biofriendly alternative to Xylene: a chemical solvent with carcinogenic properties that is used in histological staining procedures. To give some background information, Xylene is used as a clearing agent to clarify tissue samples that were embedded in paraffin wax. These deparaffinization steps are done after the wax-embedded tissue sample has been cut using a microtome for tissue sectioning. This allows me to obtain extremely thin tissue samples. After a successful experiment of replacing Xylene with pine oil in the deparaffinization steps for a Hematoxylin & Eosin staining (H&E), I’m now focused on applying pine oil as the Xylene replacement for a Masson’s Trichrome stain. Pine oil is also more cost effective than xylene since its a chemical that must be manufactured. H&E stains focus on distinguishing cellular structures in a tissue sample while Masson’s Trichrome highlights collagen and muscle fibers in tissue samples. The H&E experiment was done by previous students under my PhD mentor, Dr. Visbal. My research will be to continuing this experimental progress when I can test pine oil as a Xylene replacement during a Masson’s Trichrome stain. I am continuing to learn valuable histological techniques such as utilizing the microtome, preparing tissue samples, histological staining, and proper use of the microscope to explore cellular structures and quality of stains. Being a Hispanic woman in STEM and working with another Hispanic woman in the field is an opportunity that I am appreciative to have as it inspired to obtain my PhD and pursue becoming a scientific researcher.
Technologies
The technology used for this research was a microtome that allows for tissue sectioning for examination under the microscope. The wax-embedded tissue cube must be placed onto ince prior to cutting to obtain extremely thin pieces of mammary gland tissue obtained from mice. The samples must be cut properly to achieve optimal staining as the cutting can affect the image quality when the sample is viewed. This is a tool that is very important to science and microscopy. I have also practiced transferring and collecting the cut tissue samples by placing them in a warm water bath. This allows for the wax to be smoothed out before collecting the sample on a slide since they are embedded in paraffin wax. Samples on the slides are then dried to be prepared for staining. Pine oil would then be used to remove paraffin wax instead of xylene. My lab partner and I came up with a Masson’s Trichrome rubric that was used to give histological scoring on previously stained mouse mammary gland tissue with Masson’s Trichrome. This rubric was created to compare scorings of cells cleared with Xylene and cells cleared with pine oil.
Explain project results
The results that came from the previous H&E staining proved that pine oil could potentially be effective in using as a clearing agent for tissue staining. The pine oil also preserved the tissue sample months after the staining which was also impressive. The goal is to find a suitable replacement that has the same preservation and clearing properties as xylene to exclude it from such procedures in the future. There is potential that pine oil could be used in the deparaffinization stage for other histological staining. If it’s proven to truly be an effective replacement, it’s a change that can be implemented globally. Hearing the work my PhD mentor did inspired me to continue the research and apply for undergraduate research programs. I want to increase the representation of Hispanic women in STEM and not let language barriers or cultural differences hold me back from opportunities.
Why it should be considered best practice?
Xylene is a biohazard that causes irritation to most of the body and could cause death at immense exposure. Xylene is also a carcinogen and constant exposure over time can lead to increased risks of a cancer diagnosis. Alternatives to Xylene in histological staining procedures can improve the safety of technicians not just at the University of Houston-Downtown but at labs and institutions around the globe. This also included less carcinogenic exposure to lab technicians and overall less chemical manufacturing for Xylene. If the research shows that pine oil is a stable replacement in preserving samples over time, then it would be more cost effective and introduces new ideas on using pine oil as a clearing agent in other histological stainings besides the two mentioned here and the possibility of testing other bio-friendly oils.
Highlights of your proposed presentation
Things I would like to highlight during the presentation is how Xylene is a carcinogen and work is being done to replace this compound for deparaffinization process in staining procedures. This raises the question in how necessary is this compounded needed for tissue staining if another natural alternative could provide the same results? I would highlight the success that the H&E stain showed using pine oil as a clearing agent for mice mammary tissue samples. I want to discuss why it is important and the benefits of possibly implementing this idea of using pine oil in other labs and institution for health concerns and reduced cost reasons.
The Evaluation Committee will evaluate submitted proposals based on the following criteria. Each area will be rated on a scale from 1 to 7 (1= non-satisfactory; 7 =outstanding), for a maximum of 63 points.