Endotoxin Testing
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- เผยแพร่เมื่อ 16 ม.ค. 2020
- Did you know that a compounded preparation that passes a sterility test is not necessarily guaranteed to be free of endotoxins? Due to the potential effects on human and animal health, it is essential to make sure that your product is endotoxin-free through the USP 85 Bacterial Endotoxin Test. Learn more about what the new regulations say regarding endotoxin testing from Eagle’s microbiologists in this video.
Related and Additional Resources:
🔬🧫 Bacterial Endotoxin Test: eagleanalytical.com/bacterial...
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#EndotoxinTesting #CompoundingPharmacy #microbiologist #USP85 #usp #fda #bet
Nice Video, was a great help for my exam. Just out of curiosity, is the quantitative analysis of endotoxine content conducted via indication/color change in the LAL test or is it a semiquantitative method using dilutions?
USP currently provides methods for three approaches to bacterial endotoxins testing when method suitability has demonstrated there is no interference.
• The gel-clot assay is commonly used as a “limit” test were samples are diluted beyond any inhibitory effect but within the limit of detection for the specific lysate and the acceptance criteria for the product. A solution of reagent in a test tube with product will ‘clot’ when the endotoxins content reaches or exceeds the lysates limit of detection. Failure to clot at the dilution used at the “product’s limit” (often about 10- or 20-fold less than the product criterion) demonstrates the product is less contaminated that the allowed content.
• A turbidimetric assay may also be used that works with a spectrophotometer that quantitatively calculates the endotoxins content based on a rate that turbidity forms due to clotting.
• Similarly, a chromogenic assay is available that releases a chromophore at a rate that allows estimation of endotoxins in the presence of a labeled lysate. The rate of color formation is measured by a spectrophotometer and is compared to a standard curve used to estimate the actual endotoxins content.
The qualitative gel-clot method may be used as a quantitative assay by assaying many dilutions of the sample across the range of detection and determining the endotoxins endpoint dilution that forms a gel. The lysate sensitivity is the endpoint, which is multiplied by the product dilution to estimate the endotoxins content. This is repeated over several replicates (similar to a most probable number test) and the geometric mean of the dilutions at the endpoints is calculated as the estimate of the of the endotoxins content.
In situations where the results of these methods differ, the results of the gel clot assay are considered official and will be the legal referee result. Spectrophotometric methods are much simpler for quantitative estimates.
This might be a helpful link: www.qgdigitalpublishing.com/publication/?i=663392&p=&pn=76
If you would like more information, feel free to email us at info@eagleanalytical.com
Please do you know the most recent endotoxin test. There's a new one that's not BET or pyrogen rabbit test. Thanks
Thank you for the very informative video. I have the opposite question. Does the failure of BET or endotoxin test signify that it is failing in sterility too?
You are very welcome and happy that you found the video informative. 😊
The answer to your question is no, you can still be sterile and fail endotoxin.
recombinant lal??
What do If I get a false positive result despite my NWC is not forming the gel
Hi @harrisonmary6552, thank you for your post. In order for us to be able to answer your question, how do you know that it is a false positive and not a true positive?
1 milion dolar question (for me very important) : endotoxin level could increase by cell culture ( multiplication) ? Or, being about dead bacteria , their substance could not be multiplied by cell culture. Someone who works in this field. Many thanks.
Endotoxins are found in the cell wall of gram-negative bacteria. The endotoxins are released in the product, or in this case, cell culture during times of cell death and when the bacterial cells are undergoing log phase (e.g. cells are actively growing and dividing). The ways to prevent endotoxin contamination is to purchase consumables (e.g. pipette tips, conical tubes, etc.) and media that are indicated as “Endotoxin-free” or “Pyrogen-free.” When marketed with this wording, the products under go a QC (Quality Control) test for the presence of Endotoxins to verify the product meets the acceptable level. If the cell culture is found to be contaminated with endotoxins, there are two options: throw out all the media, reagents, and consumables or introduce a reagent that can eliminate the lipopolysaccharide protein from the cell culture. An example of this is Triton X-114.
@@eagleanalytical5957 Many thanks.
@@funny11744 You're very welcome.
the answer to your question is : YES
@@szilard16 thank you
sterility test is not necessarily guaranteed to be free of endotoxins? Due to the potential effects on human and animal health, it is essential to make sure that your product is endotoxin-free through the USP 85
It is important to reiterate that endotoxins are released from the cell wall of gram-negative bacteria during the sterilization process. Endotoxin testing is a non-viable test. Sterility testing is testing for the presence of *viable* microorganisms. This is why you can have a sample pass endotoxins but fail sterility.
I WANT TO SEE STEP BY STEP PROCESS NOT A OVERALL
Why does the guy have the mask under his nose in 1:25 seconds??
That microbiologist is wearing a beard cover, also known as a beard net. Beard covers/nets are often worn in laboratory or cleanroom environments and are a simple yet effective way to prevent beard and other facial hair from shedding contamination.