Mycoplasma Screening

Bioresearch Support Core

Overview

Mycoplasma infection in cell culture would alter not only cell behavior but also genetic content in the cell, hence compromising both phenotypic and genotypic experimental results. Being a major concern in cell culture laboratories, the Core have been providing weekly free mycoplasma PCR detection service for research labs in Faculty of Medicine since 2018. As the mycoplasma is easy to transmit but difficult to eradicate, regular contamination checking for the cell culture and proper handling of the contaminated culture are highly recommended.

Sample Submission

Sample Preparation Guide

Various form of samples can be prepared and submitted as the guide below.

*Each PI group could submit maximum of 20 samples per week

*DO NOT submit pathogen / pathogen-infected sample or any sample of biosafety level 3

A. Cell Culture Medium

A1. Cells should be cultured in antibiotics-free medium for at least 3 days (Over 1 week is preferred). Do not change medium for at least 3 days prior to sample collection.

A2. Collect a total of 3ml medium in two 2 ml microcentrifuge tubes (i.e. 1.5 ml in each 2 tube; sample in cryo-tube will NOT be accepted). 

Autoclaved 2 ml centrifuge tubes are available from the Core at L6-01, Lab Block  if you do not have them in your own lab. Original pack (Axygen MCT-200-C) could be purchased from Online Ordering system in Bioreagent Core for frequently sample preparation.

A3. Store the sample tubes at 4 oC until submission.

B. Extracted DNA

B1. Aliquot at least 100ul of extracted DNA in one 1.5ml micro-centrifuge tube. Concentration of the host DNA is not concerned.

B2. Store the sample tubes at -20 oC until submission.

C. Surface Swab

C1. Swab can be used to check the presence of mycoplasma on working surfaces (e.g. incubator, BSC). Swabs are available from the Core at L6-01, Lab Block.

C2. Swab the surface against the area of interest and put the swab back into the sample tube with 3ml PBS.

C3.  Store the samples at 4 oC until submission.

4. Label your sample tubes clearly with “Initial of your name – sample number” on the tubes. The tubes for the same sample shoulf be labelled with the same name.
    e.g. For CHAN, Tai Ming with 3 samples. Please label the tubes as: CTM-1, CTM-2, CTM-3 (each in duplicate).

5. Fill in the online sample submission form at: https://goo.gl/forms/sH7FeAfuCRa7zvcp1. A confirmation email will be generated after submission.

6. Pack and submit your sample(s) with a paper note with “User Name” and “PI Name” in a sealed zipper bag to the sample submission ice box placed near L6-01, Laboratory block, and fill in the log book properly. Any sample submitted without the note with “User Name” and “PI Name” in the same bag will NOT be processed!

Schedule

Samples can be submitted to L6-01, Laboratory block during 10am-5pm every Tuesday. (Submission date will be changed to the followed Wednesday if Tuesday is a public/university holiday. No test will be run in the week with holiday on Wednesday).

Detailed schedule can be reviewed from the calendar at:
https://calendar.google.com/calendar/embed?src=mycoplasmafcf%40gmail.com&ctz=Asia%2FHong_Kong

Any enquiry, please contact corefac@hku.hk or 3917 9440.

Decontamination

For cell culture found to be contaminated with mycoplasma, it is strongly recommended to replace and discard/autoclave all culture/stock. In exceptional case where the contaminated line is irreplaceable, antibiotic treatment may be considered for decontaminating the culture. However, success of decontamination is not guaranteed due to resistance of some mycoplasma strains and loss of the culture due to cytotoxic effects of the antibiotic.

Unless it is absolutely essential, the infected cultures should be discarded. If decontamination is the only option, antibiotics treated samples are requested to be resubmitted for screening again (just after the antibiotic treatment and in subsequent months) to confirm the effoectiveness of the decontamination treatment. Limited amount of antibiotic Plasmocin (from Invivogen) for treatment is available from the Core on request. Decontamination reagent request Form should be completed and submitted to corefac@hku.hk or L6-01, Lab Block. Additional amount of antibiotic Plasmocin may also be purchased through Bioreagent Core Online Ordering System.

Contact us for other alternative if treatment with Plamocin is not sucessful,.

 

References:Mycoplasma_Screening_References.pdf

For any enquiries, please contact corefac@hku.hk or 3917 9440.

@2021 Centre for PanorOmic Sciences (CPOS), Li Ka Shing Faculty of Medicine, The University of Hong Kong. All rights reserved.