Endotoxins are of great concern in the medical device and pharmaceutical industries and, due to the implications of their presence, are now of growing concern in the biotech and research fields. Endotoxins are a pyrogen, which is defined as a substance that can cause a fever response. They are also toxic to cells grown in tissue culture conditions. Endotoxin contamination can affect general laboratory consumables. If the product is not certified to be free of endotoxins, many experiments may fail.
Endotoxins are found in the lipopolysaccharide complex of the outer membrane of Gram negative bacteria. When the cell dies, the membrane of the cell ruptures and endotoxins are released into the surrounding environment. This can cause contamination that may have adverse effects on a variety of applications and experiments. Endotoxins can be found in the air, water, soil, via human contact, and any non-sterile environment.
Certifying products endotoxin free has become a common practice among general laboratory plastics manufacturers as well as medical and drug manufacturers. This is due to the ability of endotoxins to interfere with many common laboratory techniques such as: the creation and maintenance of cell culture lines, and the transfection of cell lines with molecular reagents. The presence of endotoxins is an indication of the overall purity and microbial activity of a particular product or raw material. Testing your product lines for this common contaminant will add integrity to your products and will allow a wider range of customers to utilize your products for their experiments and applications.
MO BIO Laboratories, Inc. and all its testing technicians have been qualified according to FDA regulations to utilize the LAL (Limulus Amebocyte Lysate) Gel Clot method to test raw materials or end products for the presence of endotoxins. This method uses components found in the blood of the Blue Horseshoe Crab, Limulus polyphemus, which forms a gel-like clot when incubated in the presence of endotoxins. This method is used to determine if products or materials are "endotoxin free". The sensitivity or detection limit of the endotoxin test is 0.06 EU/ml (Endotoxin Units). Products and materials can be certified to a sensitivity of 0.03 EU/ml upon request. Notice of at least 3 business days needs to be received by the Services Department upon requesting this service.
Upon receiving the product to be tested at Mo Bio Laboratories, a product extract is made and then exposed to and incubated with the horseshoe crab lysate (LAL). The product extract is incubated along with a standard series of the Control Standard Endotoxin (CSE) as the positive control and the unexposed extract fluid as a negative control. After the incubation period, the tubes containing the controls and the extract are observed for the presence of the gel clot. A clot indicates the presence of endotoxins. No clot indicates absence of endotoxins (i.e. < 0.06 EU/ml).
Once the certification assay is completed, the customer is contacted with the preliminary results if indicated and a report containing a Data and Results sheet and a Certificate of Analysis is mailed to the customer.
When using the LAL Gel Clot method, there are two separate types of tests that can be performed on a particular product, a qualtitative or a quantitative assay.
The quantitative LAL assay is usually used on liquid products in which a Preliminary Assay and a Validation of a dilution series is performed. ALthough not common, the quantitative assay can be performed on devices. The Preliminary Assay tests for the pH of the product. The pH must be compatible with the LAL gel clot assay in order for a validation to be performed. This Validation tests the product dilution series for an endpoint which is used to calculate the final range of endotoxin concentration present in the product.
Qualitative testing also utilizes the LAL Gel Clot method but rather than trying to detect the quantitative level of endotoxins, it tests the endotoxin levels of the product to be either greater than or less than the sensitivity of the test, which is 0.06 EU/ml. This type of test can be performed on solid and liquid products.
To begin certifying your product Endotoxin-Free, a sample of the product must be received by our Services Division to evaluate how best to test your particular product. Preliminary testing will be required for any raw material or liquid product. Preliminary testing will provide the concentration at which your product can be routinely tested. An extraction procedure must be determined for most devices. Once an extraction procedure for a device is approved, it is used each time that product is received for testing. An Inhibition/Enhancement Validation must be performed before Routine Testing can be performed on any product.
Many products will fit one of our pre-designed extraction procedures, such as microcentrifuge tubes, PCR tubes and pipet tips. However, because all products are not the same, each product received is evaluated on an individual basis. Liquid solutions, reagents and solids, due to the diversity of these products, require a separate test procedure to be designed each time a product of this type is submitted.
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You and your company decide upon a testing schedule that best fits the needs of your particular product. MO BIO Laboratories, Inc. can work with you to design a testing schedule that accomplishes this goal. Click here for more information.
Download an Excel file to submit your samples for testing by MO BIO.