Preventing Contamination In Cell Culture, Day 32

Cell culture is the beating heart of experimental immunology.  

𝗦𝗼𝘂𝗿𝗰𝗲𝘀 𝗼𝗳 𝗖𝗼𝗻𝘁𝗮𝗺𝗶𝗻𝗮𝘁𝗶𝗼𝗻 
Human: skin, hair, clothing fibers. 
Consumables: pipettes, plates, or flasks, not properly sterilized. 
Reagents: serum, cytokines, media supplements not sterile-filtered. 
Environment: dust, aerosols, incubator condensation [1]. 

𝗕𝗲𝘀𝘁 𝗣𝗿𝗮𝗰𝘁𝗶𝗰𝗲𝘀 𝗨𝗻𝗱𝗲𝗿 𝘁𝗵𝗲 𝗛𝗼𝗼𝗱 
The biosafety cabinet (BSC) maintains laminar airflow, protecting both the scientist and the cells. 

𝗢𝗯𝘀𝗲𝗿𝘃𝗮𝘁𝗶𝗼𝗻𝘀 𝗮𝗻𝗱 𝗗𝗶𝗹𝗲𝗺𝗺𝗮𝘀 𝗳𝗿𝗼𝗺 𝘁𝗵𝗲 𝗟𝗮𝗯 
𝗗𝗶𝗹𝗲𝗺𝗺𝗮 𝟭 – 𝗣𝗶𝗽𝗲𝘁𝘁𝗲 𝗣𝗮𝗰𝗸𝗮𝗴𝗲𝘀 
In some labs, unopened stripette packages are stored inside the hood and sprayed with 70% isopropanol before opening. While intended to maintain sterility, it raises a question: why place the sealed package inside at all? The exterior of the package may carry microbes, but over-spraying can create extra liquid or aerosol. 
𝗕𝗲𝘀𝘁 𝗽𝗿𝗮𝗰𝘁𝗶𝗰𝗲: only bring what you need, open as close as possible to the airflow, and avoid clutter.  

𝗗𝗶𝗹𝗲𝗺𝗺𝗮 𝟮 – 𝗠𝗲𝗱𝗶𝗮 𝗳𝗿𝗼𝗺 𝗖𝗼𝗹𝗱 𝗦𝘁𝗼𝗿𝗮𝗴𝗲 
Researchers often take media or serum from the fridge and insert it directly under the hood, or thaw supplements in a water bath and place them immediately inside. Some spray bottles, some do not. 
𝗕𝗲𝘀𝘁 𝗽𝗿𝗮𝗰𝘁𝗶𝗰𝗲: always wipe bottles with 70% isopropanol before entering the BSC.  

𝗔𝗻𝗲𝗰𝗱𝗼𝘁𝗲 – 𝗧𝗵𝗲 𝗚𝗹𝘂𝗲 𝗜𝗻𝗰𝗶𝗱𝗲𝗻𝘁 
During my Master’s thesis, I once used office glue from Tedi to attach explanted rodent aortic tissue onto a culture plate. I wore gloves, but the glue was not sterile. The samples went into the incubator and, the next day, the medium turned cloudy and milky – classic bacterial contamination. My supervisor explained that UV sterilization or medical-grade adhesives like Dermabond should have been used. 
𝗟𝗲𝘀𝘀𝗼𝗻 𝗹𝗲𝗮𝗿𝗻𝗲𝗱: sterility cannot be improvised. What goes into the hood must already be clean. 

𝗪𝗵𝘆 𝗜𝘁 𝗠𝗮𝘁𝘁𝗲𝗿𝘀 
Contamination alters experimental outcomes: 
A. Bacteria and fungi secrete metabolites affecting cytokine responses. 
B. Endotoxin activates immune cells in unintended ways [2]. 
C. Human contaminants (skin, hair) introduce foreign DNA and proteins [3]. 
Even small contamination events can invalidate months of immunology experiments.  

𝗤𝘂𝗲𝘀𝘁𝗶𝗼𝗻 𝗳𝗼𝗿 𝘁𝗵𝗲 𝗮𝘂𝗱𝗶𝗲𝗻𝗰𝗲:  What is the most unforgettable contamination lesson you’ve experienced in your lab?  

Stay tuned for 𝗗𝗮𝘆 𝟯𝟯: 𝗨𝗻𝗱𝗲𝗿𝘀𝘁𝗮𝗻𝗱𝗶𝗻𝗴 𝗔𝗱𝗵𝗲𝗿𝗲𝗻𝘁 𝗮𝗻𝗱 𝗦𝘂𝘀𝗽𝗲𝗻𝘀𝗶𝗼𝗻 𝗖𝗲𝗹𝗹 𝗖𝘂𝗹𝘁𝘂𝗿𝗲𝘀 

𝗥𝗲𝗳𝗲𝗿𝗲𝗻𝗰𝗲𝘀 
1. doi: 10.1023/A:1022913015916 
2. DOI: 10.1126/science.2402637 
3. DOI:10.1002/9780470649367   

hashtag#100DaysOfImmunology hashtag#CellCulture hashtag#Contamination hashtag#LabSafety hashtag#Biosafety hashtag#Sterility hashtag#ImmunologyResearch hashtag#iPSC hashtag#CAR_T hashtag#LaboratoryPractice