Sartorius Microbiology Products Keeping your Quality under Control

Transcription

1 Sartorius Microbiology Products Keeping your Quality under Control Introducing: Real-time PCR Mycoplasma Detection Kit for Universities & R&D Labs Microsart RESEARCH Mycoplasma

2 Table of Contents A. laidlawii colony after 5 days of incubation, 120x magnification Introduction Markets & Customers Target Application Product Information Features & Benefits Ordering & Price Information Competition Frequently asked questions Literature & Documentation Page 2

3 We completed now our Mycoplasma Product Family Microsart AMP Mycoplasma Microsart RESEARCH Mycoplasma Microsart ATMP Mycoplasma

4 Complete range for different customer needs Real-Time PCR Kits for Sensitive Mycoplasma Contamination Control for All Safety Levels Microsart AMP Mycoplasma Microsart ATMP Mycoplasma Microsart RESEARCH Regulated in-process and lotrelease testing, full EP compliance Highest sensitivity through large volume Validation needed Highly reliable Testing of autologous cell transplants (ATMPs like chondrocyte cultures) Cost efficient testing Easy handling Process control Specific validation needed Testing of cell culture materials in research and development Easy hands -on Fast Cost saving Screening small sample volume Page 4

5 In a Nutshell... What are Mycoplasma? Smallest self-reproducing bacteria: 0.2 to 0.8 μm No cell wall Easily pass through 0.45 μm filters, and even through sterilizing grade filters with pore sizes of 0.2 µm Page 5

6 Why it is so important to control the cell culture! Approx. 5-85% of all continuous human or animal cell lines are contaminated with mycoplasmas. (source: Laborjournal) Contamination might have no or no obvious effects A contamination can change the properties of the cells & could destroy all results of comprehensive R&D phases Page 6

7 Possible sources of Mycoplasma contamination Primary cultures from the original tissue (incidence approximately 4 %) Cross contamination contaminated cultures new cultures from unknown sources, also partly from cell banks virus suspensions, antibody- solutions or other additions of contaminated cell cultures Direct contamination serum (only treated serums, e.g. UV or γ-radiated are presumably Mycoplasma free) laboratory instruments, media and reagents, which came into contact with contaminated cultures The User handling droplet transfer insufficient disinfection careless technical work Page 7

8 Markets and Customers The Microsart RESEARCH Mycoplasma detection kit, as the name suggests, is especially designed for all: R & D labs Governmental Research Institutes Universities All labs who want to detect mycoplasma without the need of E.P regulatory compliance Page 8

9 Target Application Mycoplasma Detection in R&D Labs Customer segment Research & Development Labs Types of Samples Cell culture media New cell lines Cell culture supernatants etc. What are the needs? Early detection of potential contamination Fast results Easy to use test procedure Reliable results What are the risks? Loosing the complete experiment Having to repeat everything Associated financial burden (time & money)

10 What s important for the customers? Customers needs Sartorius Solution Convenience Easy and fast solutions 3 steps kit Simple handling Wide compatibility with the common-used PCR cycler Quality of results Reduces the risk of false positive results false negative results Uses TaqMan technology as a probe high specificity

11 Fast time to result Optimized Workflow Time to result in less than 3 hours due to fast thermal profile and less hands-on time - Ease of use - Quick response for corrective action - Minimal operator training

12 Reduction of False Negative results Positive & Internal control reaction Positive control included in each Kit Internal inhibition control in each sample tube for monitoring the process (no occurrence of PCR inhibition)

13 Reduction of False Positive results Specific DNA amplification based on TaqMan probe Amplification is specific to Mycoplasma DNA No amplification of non-mycoplasma DNA (background DNA) Results are reliable and highly specific

14 Microsart RESEARCH Mycoplasma Product Information Page 14

15 Everything is included... Mycoplasma Mix (including probe, primers, nucleotides & polymerase) Rehydration Buffer Positive Control PCR grade Water Page 15

16 TaqMan probe vs. other technologies Microsart RESEARCH Taq-Man Probe SYBER Green Dye Traditional PCR Biochemical Test Saves valuable time due to the use of highly specific TaqMan probes, the analysis can be carried out quickly using easy to interpret fluorescence curves. Binds to any double strand DNA in the sample. After the DNA amplification a melting curve analysis has to be done which needs additional time. Unspecific amplification products such as primer artefacts make it difficult to interpret the results correctly. Sample is analyzed on an agarose gel, i.e. yes/no result. Low DNA concentration cause weak bands no clear/objective result interpretation possible. The biochemical test exploits the activity of certain mycoplasma enzymes. The viable enzymes react with the substrate and convert ADP to ATP. Increase in ATP can be detected using a luminometer.

17 Only 3 Steps for Sample Preparation and Amplification Optional 1. Step DNA Extraction up to 200 µl 1/2. Step 2 µl sample direct for PCR setup 3. Step PCR & Result Interpretation The Microsart RESEARCH Mycoplasma Detection Kit contains all necessary components (including polymerase) and shortens the total procedure - from DNA extraction to PCR result - down to a few hours. Page 17

18 Important Information storage, transportation and license Temperature sensitive! Shipping only under cold-chain transportation conditions! The cold chain may not be interrupted! Method of transport must be per courier! Limited License for sales in USA, Canada & Japan Use of this product is covered by one or more of the following US patents and corresponding patent claims outside the US: 5,804,375, 5,538,848, 5,723,591, 5,876,930, 6,030,787 and 6,258,569. The purchase of this product includes a limited, non-transferable immunity from suit under the foregoing patent claims for using only this amount of product The patent expires end of November 2015 Solely in Industrial Microbiology QA/QC Field, including reporting results of purchaser s activities for a fee or other commercial consideration, and also for the purchaser's own internal research. No right under any other patent claim is conveyed expressly, by implication, or by estoppel. Further information on purchasing licenses may be obtained from the Director of Licensing, Applied Biosystems, 850 Lincoln Centre Drive, Foster City, California 94404, USA. Page 18

19 Ordering information & pricing Page 19

20 Ordering information & pricing Order No. Description Quantity Price SMB Microsart RESEARCH Mycoplasma, Mycoplasma Real-time PCR ,- SMB Microsart RESEARCH Mycoplasma, Mycoplasma Real-time PCR ,- SMB Microsart AMP Extraction Kit ,- Page 20

21 Thermal Stability unopened and stored at +2 to +8 C: 18 months rehydrated and stored at < -18 C: 6 months rehydrated and stored at +2 to +8 C: < 8 hours rehydrated and stored at room temperature < 60 min... because fluorescence dyes are sensitive to temperature & light exposure) Page 21

22 Competitors

23 Fast time to result Microsart RESEARCH Mycoplasma MycoSEQ (Life) Time to result about 3 hours only due to fast thermal profile and few required steps - Quick response for corrective action - Efficient handling Time to result about 6 hours due to longer thermal profile and longer sample preparation protocol - Slower response for corrective action

24 Save hands-on time & Easy interpretation Microsart RESEARCH Mycoplasma MycoTool (Roche) Very cost efficient (9 /per test) Most expensive (500 /per test) Very easy set up very convenient No specific equipment is needed (any Real-time cycler can be used) Real-time PCR quantification possible Extensive hands-on time!!! Complicated DNA extraction Polyacrylamide Gels often rip/toxic A lot of additional equipment necessary Conventional PCR no quantification

25 Save hands-on time & Easy interpretation Microsart RESEARCH Mycoplasma MycoAlert Plus (Lonza) Very easy set up very convenient No specific equipment is needed Internal control no additional control reaction No limitation in parallel testing; run as many reactions in parallel as the thermal cycler allows Sensitivity is less then 50 CFU/ml Luminometer is needed No internal control Limited to 2 luminescence readings in parallel Positive control must be ordered separately

26 Save hands-on time & Easy interpretation Microsart RESEARCH Mycoplasma MycoTool RT-PCR (Roche) Very cost efficient (9 /per test) Expensive (200 /per test) Very easy set up very convenient Extensive hands-on time!!! No specific equipment is needed Complicated DNA extraction Internal control no additional control reaction A lot of additional control reactions necessary (1 Kit contains material for analyzing 10 samples which corresponds to 160 PCR reactions)

27 Frequently Asked Questions Page 27

28 To identify potential customers... watch out for this products Real-time cycler Real-time cycler PCR cooler Real-time cycler Labtop Mini-cooler PCR Hood Real-time cycler Page 28

29 Qualifying Questions Which method do you use? Are you still using the traditional methods? Do you perform Mycoplasma testing? Our Rapid Mycoplasma Detection Kits are very simple in handling Fast results using Sartorius it will only take 3 hours compared to 28 days More convenient & much more cost effective

30 FAQ What are the test principles? The kit uses the Real-time PCR technology. This molecular biological method is used to amplify and detect defined DNA sequences. The amplification is done in multiple cycles. For detection, a fluorescent-labeled probe is used. During each cycle a fluorescence measurement is performed. A logarithmic increase in signal intensity indicates a positive signal. Which target is used for amplification? The kit specifically detects the 16 S rrna genome region. How can I exclude that a negative results does not occurred due to inhibition caused by matrix effects? The kit contains an internal inhibition control DNA. Page 30

31 FAQ (cont.) Do I have to use a specific qpcr cycler? No. All Real-time cyclers which can detect FAM - and ROX labeled probes can be used with the kit. Do I need a special software for the evaluation of the results? No. The Microsart RESEARCH manual gives clear guidelines how the cycler software is used for evaluation. Page 31

32 Literature & Documentation Datasheet Flashcard Flyer Page 32

33 Thank you for Your Attention! Page 33