Kristen Krebs – Independent Project

Kristen Krebs – Independent Project

Class of 2017

Introduction to Topic

We do not currently have a method to combat sulfur mustard burns, and we cannot conduct research because no animals have skin that will react the same way as human skin. If there were an in vitro model to test on, it could be used to develop a treatment or even replaced damaged skin. While there are sure to be problems in creating this resource, it could be extremely costly to make easily avoidable mistakes. With easily accessible and simple to understand checklists, valuable time and money could be saved.

Project Description

I spent eight weeks at the United States Army Medical Research Institute of Chemical Defense (USAMRICD) under the mentorship of Erik Eaton and Timothy Varney, PhD. Sulfur mustard is a current threat to American soldiers in the Middle East and potentially even to people here in the U.S. Our goal was to develop an in vitro human skin model for research purposes to combat the hazardous gas. During my time there, I learned the basic procedures for culturing cells and have noticed some potential slip-up points in the process.

The lab procedures we used became so routine that once we learned them, we did not need a mentor hovering over our shoulder when completing them. This also could be dangerous as it is easy to make a simple mistake, such as missing one small step that hinders the project from progressing. I have developed a series of checklists using Trello to follow when coating flasks, plating cells, feeding cells, and passing cells. While there was an official procedure typed up for the specific project being worked on, it was not very detailed and often not even referred to. Having the checklist on Trello allows it to be easily accessed from any device and ensures no steps are forgotten.

Experience Description

In order to get the internship, I had to apply through the Army Educational Outreach Program (AEOP) in February 2016. In April, I received an email from Sergeant Erik Eaton from USAMRICD who was selecting three students to work on his team. Once I was selected, I had to wait to hear back from AEOP for confirmation.

The first day, we had to sit through in processing presentations to learn about safety and standard procedures. Then we met our mentors and started learning about our projects. Since there were three students on my team, each of us took a layer of the skin to work on for our in vitro model. I had the epidermis, so my job was to bring up keratinocytes and eventually add them to the other layers of the skin.

Week one in the lab was spent getting adjusted to all of the new equipment and procedures we needed to complete. We started by coating flasks and plating the cells. The most difficult part was remembering how to use the equipment and complete the tasks without messing something up. When using the pipette controller, it was important to hold level so that the liquid would not get into the filter. The tip of the pipette also could not touch anything or it would become unsterile. Trying to balance and remember that while moving something with the other hand was challenging. Once I had completed some of these tasks a few times and I had become used to them, it became easier to pick up new ones. Feeding the cells the cells and eventually passing them were just different combinations of steps I already knew. Over time, we continued to grow the cells, but my keratinocytes were not growing well. I restarted with a new batch which grew better, but still not as well as they should have. Towards the end, we started to combine all of the different cells to make the layers of the skin. Human mesenchymal stem cells (HMSC) were plated in the bottom of well insert plates to differentiate into the adipose of the hypodermis. A gel was made of collagen and fibroblasts to be the dermis and was placed on the insert, and my keratinocytes were put on top of that layer. Due to time constraints, we were unable to get any further, but our mentors have continued to work on the project.

From my experience, I have learned a lot about how professional labs function, and I have a great appreciation for people who devote their time to research. I do not think I would want to work in research because I never realized all of the desk work that is also involved with it. I only envisioned that lab work but failed to realize they have to learn what research already exists which could be boring and tedious at times.

Innovation Description

After a couple of weeks of being in the lab, I as well as the other students, had gotten used to all of the procedures we had to complete. One day, the HMSCs had to be passed into a larger flask, so the team member in charge of those cells began to do so. Everything seemed to be normal until she started to put the cells, trypsin, and trypsin inhibitor into the conical tubes to be spun down, as there was way more liquid than there should have been. We realized that the media had not been aspirated before starting the process. The simple step had been overlooked even by those of us just watching.

Around the same time, I read The Checklist Manifesto and realized that a checklist would have most likely prevented the whole ordeal (Gawande, 2016). We did have protocol that was typed out on paper, but it was very broad and not always nearby. It included steps such as “Replace medium” and “Plate to T-180,” and the basic steps were not accounted for (Eaton, 2016). My innovation takes the concept of the protocol and breaks it up into easy to use checklists on Trello. Using Trello allows the user to pull it up on any device, and new checklists can be made specifically for a cell type or project whenever necessary.

The board I made consists of five lists. The first is a key to define any acronyms or labels used so anyone observing it can understand it. The other four lists are checklists for different tasks. The first two are coating flasks and plating cells. While they are different tasks, coating the flasks flows right into plating the cells. The other checklists are feeding cells and passing cells. These are all procedures used for basic cell culturing specific to human epidermal keratinocytes (HEK). The checklists are in a do-confirm format, so every few steps, there is a pause point (Gawande, 2010). Some are general and just make sure the previous steps have been done, and others give instructions on what to do if the previous steps have not been done correctly. All of the pause points start off labeled as red and can be changed to green once they are complete. Anything on a checklist that is extra important to pay attention to is labeled as yellow.

The overall work of this innovation adds up to over 10 hours. I spent two hours working with my mentor to review the protocol and brainstorm what should be included in the checklists (Eaton, 2016). I spent one hour reviewing the features of Trello (Cervino, 2015). I spent two hours generating the steps needed to make up the checklists, one hour placing them into the proper lists, and one hour sorting all of the proper labels as incomplete, complete, and in progress. After meeting with Mr. Burke for my final measurable objective, I spent an hour reviewing the advice given and checklist for checklists from The Checklist Manifesto (Gawande, 2010). From there, I spent two and a half hours reorganizing the cards making the steps easier to follow, changing the labels as necessary, and adding the pause points.

Project Topic

Introduction to Topic

We do not currently have a method to combat sulfur mustard burns, and we cannot conduct research because no animals have skin that will react the same way as human skin. If there were an in vitro model to test on, it could be used to develop a treatment or even replaced damaged skin. While there are sure to be problems in creating this resource, it could be extremely costly to make easily avoidable mistakes. With easily accessible and simple to understand checklists, valuable time and money could be saved.

Project Overview

Project Description

I spent eight weeks at the United States Army Medical Research Institute of Chemical Defense (USAMRICD) under the mentorship of Erik Eaton and Timothy Varney, PhD. Sulfur mustard is a current threat to American soldiers in the Middle East and potentially even to people here in the U.S. Our goal was to develop an in vitro human skin model for research purposes to combat the hazardous gas. During my time there, I learned the basic procedures for culturing cells and have noticed some potential slip-up points in the process.

The lab procedures we used became so routine that once we learned them, we did not need a mentor hovering over our shoulder when completing them. This also could be dangerous as it is easy to make a simple mistake, such as missing one small step that hinders the project from progressing. I have developed a series of checklists using Trello to follow when coating flasks, plating cells, feeding cells, and passing cells. While there was an official procedure typed up for the specific project being worked on, it was not very detailed and often not even referred to. Having the checklist on Trello allows it to be easily accessed from any device and ensures no steps are forgotten.

Experience

Experience Description

In order to get the internship, I had to apply through the Army Educational Outreach Program (AEOP) in February 2016. In April, I received an email from Sergeant Erik Eaton from USAMRICD who was selecting three students to work on his team. Once I was selected, I had to wait to hear back from AEOP for confirmation.

The first day, we had to sit through in processing presentations to learn about safety and standard procedures. Then we met our mentors and started learning about our projects. Since there were three students on my team, each of us took a layer of the skin to work on for our in vitro model. I had the epidermis, so my job was to bring up keratinocytes and eventually add them to the other layers of the skin.

Week one in the lab was spent getting adjusted to all of the new equipment and procedures we needed to complete. We started by coating flasks and plating the cells. The most difficult part was remembering how to use the equipment and complete the tasks without messing something up. When using the pipette controller, it was important to hold level so that the liquid would not get into the filter. The tip of the pipette also could not touch anything or it would become unsterile. Trying to balance and remember that while moving something with the other hand was challenging. Once I had completed some of these tasks a few times and I had become used to them, it became easier to pick up new ones. Feeding the cells the cells and eventually passing them were just different combinations of steps I already knew. Over time, we continued to grow the cells, but my keratinocytes were not growing well. I restarted with a new batch which grew better, but still not as well as they should have. Towards the end, we started to combine all of the different cells to make the layers of the skin. Human mesenchymal stem cells (HMSC) were plated in the bottom of well insert plates to differentiate into the adipose of the hypodermis. A gel was made of collagen and fibroblasts to be the dermis and was placed on the insert, and my keratinocytes were put on top of that layer. Due to time constraints, we were unable to get any further, but our mentors have continued to work on the project.

From my experience, I have learned a lot about how professional labs function, and I have a great appreciation for people who devote their time to research. I do not think I would want to work in research because I never realized all of the desk work that is also involved with it. I only envisioned that lab work but failed to realize they have to learn what research already exists which could be boring and tedious at times.

Innovation

Innovation Description

After a couple of weeks of being in the lab, I as well as the other students, had gotten used to all of the procedures we had to complete. One day, the HMSCs had to be passed into a larger flask, so the team member in charge of those cells began to do so. Everything seemed to be normal until she started to put the cells, trypsin, and trypsin inhibitor into the conical tubes to be spun down, as there was way more liquid than there should have been. We realized that the media had not been aspirated before starting the process. The simple step had been overlooked even by those of us just watching.

Around the same time, I read The Checklist Manifesto and realized that a checklist would have most likely prevented the whole ordeal (Gawande, 2016). We did have protocol that was typed out on paper, but it was very broad and not always nearby. It included steps such as “Replace medium” and “Plate to T-180,” and the basic steps were not accounted for (Eaton, 2016). My innovation takes the concept of the protocol and breaks it up into easy to use checklists on Trello. Using Trello allows the user to pull it up on any device, and new checklists can be made specifically for a cell type or project whenever necessary.

The board I made consists of five lists. The first is a key to define any acronyms or labels used so anyone observing it can understand it. The other four lists are checklists for different tasks. The first two are coating flasks and plating cells. While they are different tasks, coating the flasks flows right into plating the cells. The other checklists are feeding cells and passing cells. These are all procedures used for basic cell culturing specific to human epidermal keratinocytes (HEK). The checklists are in a do-confirm format, so every few steps, there is a pause point (Gawande, 2010). Some are general and just make sure the previous steps have been done, and others give instructions on what to do if the previous steps have not been done correctly. All of the pause points start off labeled as red and can be changed to green once they are complete. Anything on a checklist that is extra important to pay attention to is labeled as yellow.

The overall work of this innovation adds up to over 10 hours. I spent two hours working with my mentor to review the protocol and brainstorm what should be included in the checklists (Eaton, 2016). I spent one hour reviewing the features of Trello (Cervino, 2015). I spent two hours generating the steps needed to make up the checklists, one hour placing them into the proper lists, and one hour sorting all of the proper labels as incomplete, complete, and in progress. After meeting with Mr. Burke for my final measurable objective, I spent an hour reviewing the advice given and checklist for checklists from The Checklist Manifesto (Gawande, 2010). From there, I spent two and a half hours reorganizing the cards making the steps easier to follow, changing the labels as necessary, and adding the pause points.

By | 2017-05-15T15:49:21+00:00 May 15th, 2017|Biomed Capstone Project 2017|0 Comments

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