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Frequently Asked Questions

Endotoxin Testing

Questions Regarding the Endosafe®-PTS
Is the PTS licensed by the FDA?
Yes, Charles River Laboratories received FDA approval for the PTS in July 2006. With this approval, the PTS is an approved LAL testing method for in-process and end product release.
What is the QC procedure for cartridges?
The cartridges are potency tested, spike recovery is performed and the calibration code is determined. The calibration code contains all of the cartridge test parameters that were determined during potency testing, as well as the archived curve for that batch of cartridges. The calibration code is then verified with an additional test cartridge.
How is the archived standard curve determined?
Five (5) cartridges of each standard curve concentration are run to obtain the average reaction time for each standard; five (5) cartridges are used for the negative water control; ten (10) cartridges are used to determine the spike concentration; and one (1) cartridge is used for potency and calibration code verification.
What are the QC requirements for release of the test cartridges?
The archived curve is determined, as mentioned above, with acceptability requirements of <25% Coefficient of Variation between replicates and a Correlation Coefficient (R) of = 0.990. Recovery of the spiked values in the potency verification phase must be within 50% to 200%.
How is the potency of the spike determined?
LAL Reagent Water is tested on 10 test cartridges, and the mean reported value is calculated and becomes the archived spike concentration for that lot of cartridges. This value may be different for each batch of cartridges.
How is the spike recovery calculated on the PTS?

The % spike recovery can be calculated by dividing the reported spike EU/mL value by the archived spike concentration determined during potency testing and then multiplying by 100. The "Recovery Factor" displayed on the test report is the conversion factor for the actual spike concentration. It must be used to calculate the actual spike concentration for that particular lot of cartridges by multiplying it by the midpoint of the archived standard curve.  

For example:
0.577 EU/mL (Reported spike value)  > x   100 = 111.0% spike recovery
0.52 EU/mL  (Archived spike concentration)

Why is the positive product control not 0.5 EU/mL for a 5 to 0.05 EU/mL curve?
The USP says to select an endotoxin concentration at or near the middle of the endotoxin standard curve. The cartridge is spiked with a predetermined nanogram amount of CSE during the filling operation. The EU/ng value is then determined during release testing using the government Reference Standard Endotoxin (RSE).
Can the PTS data be downloaded directly to a regular printer?
Charles River offers a battery powered, rechargeable thermal printer that works directly with the reader as well as an Epson printer that prints in black ink on receipt paper for a long-lasting test record. To download data to a separate printer, we recommend using the PTS Logger software that is included with the purchase of the reader.
Can multiple tests be printed at one time?
With the PTS thermal printer or the Epson printer attached to the reader, you can print results from the last test that was run, print results by date, or print all the results stored in the reader (up to 100).
Can I export the data to Excel?
Not directly. There is the possibility that data that is downloaded from "Hyper Terminal" may be exported to Excel, but we have no experience with this.
What types of products have been tested with the PTS?
A range of products have been tested with the PTS, including biologicals (Human Serum Albumin), raw materials (saline, buffers), parenterals, water systems, and medical device extracts. Since the PTS cartridges are loaded with licensed LAL, any product that is tested by other LAL methods can be tested with the PTS.
Will the PTS read hot or cold samples?
Extreme temperature may affect results and, therefore, we recommend that you treat your PTS samples as you would any other LAL sample. Bring test sample and test cartridge to room temperature before using. Load the cartridge into the reader and wait until "Add sample" is on the display before delivering sample to cartridge.
How does the cost compare to other LAL methods?
At first glance, it may appear that the cartridges are more expensive than other quantitative LAL methods. Keep in mind, however, that all the reagents needed to run the test - lysate, endotoxin, and LRW - are already included in the cartridge. The standard curve has also been performed already, which typically requires a minimum of 6 wells and 2 additional wells for negative water controls. So one test run with the PTS really includes 8 wells for the curve and 4 wells for the samples and negative controls - a total of 12 wells. The advanced technology of the test also saves considerable time on labor as well.
Do you have a cartridge that has a lower sensitivity (0.01 EU)?
Yes, the lowest sensitivity now available is 0.01 EU/mL.
What is the maximum time setting for the assay to complete?
The assay time is dependent upon the specific lot of cartridges and their sensitivity. The time is determined by the calibration code and can not be set by the user. A typical assay is takes approximately 15-20 minutes to complete.
Will you have a system that will fit more than one cartridge?
We are considering many different configurations for expanded models of the PTS.
How can I calibrate my instrument on a regular basis? How often should I?
The factory calibration certified at the time of initial shipment is valid for one year. Re-certification cannot be done by a customer, so Charles River offers a calibration re-certification service to make sure the reader is performing as expected.
How precise and accurate is the test?
The PTS meets the same accuracy parameters as an LAL test method.
Is data generated from the PTS comparable to kinetic or gel clot LAL assays?
Yes. The PTS is a modified 2-step kinetic assay that utilizes FDA-licensed LAL reagent. Charles River is using the same government reference standard in potency testing to determine the calibration code. The same standards are used to release kinetic and gel clot LAL.
What is the expiration of the cartridges?
The current expiration period is one (1) year.
How can you tell how much charge the battery has left?
It is important to keep track of the battery life so that it does not expire unexpectedly. Charging it overnight should prevent a loss of power. When the reader is turned on, the battery will display the % charge it holds. If the reader is plugged into the charger and then turned on, it will display Bat: Charging with a percentage that slowly increases with time. If it is not plugged into the charger, it will display Bat:Discharging, and the percentage will slowly decrease with use. To view this % charge at any time, select Menu 6 to reset the reader and the % charge will be displayed. We do have plans to add a power bar to the screen to provide a constant visual reminder.
Do I need to calibrate the mini-pipettor? Does it come calibrated?
There is no need to calibrate the mini-pipettor if your QA group accepts the Certificate of Quality that verifies calibration. The manufacturer specifies that the mini-pipettor performs at ± 5% accuracy and ± 3% precision, and this document can be furnished upon request.
What is the warranty and maintenance program?
Upon purchase, the PTS comes with a warranty that is good for one year. Repairs will be made at no charge during that period. If desired, there is also an Extended Warranty program that must be purchased within the first year of ownership and will include a warranty for 4 additional years after the initial warranty has expired.
General Questions Regarding Endotoxin Testing
Is Charles River new to endotoxin testing?
Charles River Endosafe has been in the endotoxin testing business since it was founded by Dr. James F. Cooper in 1987. Dr. Cooper is one of the pioneers in the LAL field and has helped Endosafe become the worldwide market leader.
Are your reagents FDA approved?  
All Endosafe lysate, as well as the facility they are produced in, are FDA licensed.
What is your shipping policy?
Charles River ships all orders Monday through Thursday to arrive to the customer the following day.
Do you have products for oven validation?
Charles River offers the most extensive line of products for oven validation in the industry. Our endotoxin indicators are sold in 6 different concentrations ranging from 2,000 to 10 million EU / vial.
Do you sell your own automated LAL system?
Charles River sells a system that consists of an incubating plate reader along with Endosafe's endotoxin measuring and database trending software package. The system comes fully qualified along with a manual for easy in-house requalification.
Is the Endosafe software validated?
Our EndoScan-V is the first endotoxin-specific software that addresses the 21 CFR Part 11 requirement for validation. Both EndoScan-V and the BioTrend 3.0b database trending software are verified and validated consistent with USFDA and CE mark requirements and comply with CFR 820.30.
Are the horseshoe crabs killed or injured during the harvest process?
The horseshoe crabs are used during harvesting by puncturing the septum on the underside of the animal. The animal will only give as much blood as it can afford (about 25%) before it shuts off the blood supply. This ensures that the animal will not bleed to death. The horseshoe crab is then released back into the water. The mortality rate for the crab as a result of the process is less than 10%.
Why does your lysate have different properties than the competition?
Although all LAL reagents utilize the enzyme found naturally in the horseshoe crab to detect endotoxin, it is the total LAL reagent formulation that controls its performance. Such qualities as stability, sensitivity, speed and linearity are all dependent upon the individual manufacturer's LAL formulation. This formulation also controls inhibition / enhancement properties. Since its inception, Endosafe has been dedicated to developing LAL reagents that are superior to those offere by competitors.
What are the best ways to overcome interference problems?
Inhibition can best be overcome by using an LAL reagent that is designed for testing complex pharmaceutical products and raw materials. Endosafe lysate is the most recently approved in the industry, allowing us to improve upon previous interference problems.
Do you have a method to trend data from the kinetic test?  
Yes, Endosafe now offers Microtrend, a new database trending package that allows LAL customers to track and trend the various data that is generated and automatically downloaded from the EndoScan-V endotoxin measuring software.
Is chromogenic better than turbidimetric for LAL testing?
It is a misconception that chromogenic LAL is better than turbidimetric LAL. Chromogenic LAL became popular because its high sensitivity (.005 EU/ml) and 1:1 product to lysate ratio allows users to dilute further to overcome inhibition. Recent innovations by Endosafe have produced a turbidimetric lysate with the properties of a chromogenic reagent with a turbidimetric price.
What kind of technical assistance does Endosafe offer to customers?
Endosafe provides technical support via phone (800-762-7016), email, in-house troubleshooting, seminars, and workshops on a variety of topics such as methods development, SOPs, and regulatory guidance.
Does Endosafe distribute internationally?
Endosafe products are available internationally through a collection of distributors. To get a list of these distributors, click here.

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Article Highlights Success of Horseshoe Crab Management Plan

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