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konkous ☆ Greece, 2012-06-12 09:03 (4734 d 22:44 ago) Posting: # 8691 Views: 6,015 |
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Is there a specific procedure describing how whole blood samples should be treated during the interval between sampling and centrifugation? I'm referring to a BE study and to be more specific i need to know if there are any guidelines describing temperature conditions or some kind of shaking of blood samples in the vacutainers before they are centrifuged. For example if someone claims that blood samples remain at room temperature for 20 min before being centrifuged should he expect that an auditor would raise stability issues? Thank you Constantinos |
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ElMaestro ★★★ Denmark, 2012-06-12 14:51 (4734 d 16:56 ago) @ konkous Posting: # 8706 Views: 5,255 |
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Hi Konkous, ❝ Is there a specific procedure describing how whole blood samples should be treated during the interval between sampling and centrifugation? I'm referring to a BE study and to be more specific i need to know if there are any guidelines describing temperature conditions or some kind of shaking of blood samples in the vacutainers before they are centrifuged. For example if someone claims that blood samples remain at room temperature for 20 min before being centrifuged should he expect that an auditor would raise stability issues? As I see it, this should all be detailed out in your own analytical protocol / method description. You can chose the conditions under which the assay is being run and to which the validation as well as the study adheres. — Pass or fail! ElMaestro |
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Ohlbe ★★★ France, 2012-06-12 18:49 (4734 d 12:58 ago) @ ElMaestro Posting: # 8712 Views: 5,245 |
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Dear El Maestro and Constantinos, Agreed. I would suggest you to also have a look at the EU BMV guideline, end of section 4.1.9: Sufficient attention should be paid to the stability of the analyte in the sampled matrix directly after blood sampling of subjects and further preparation before storage, to ensure that the obtained concentrations by the analytical method reflect the concentrations of the analyte in the subject at the moment of sampling. A demonstration of this stability may be needed on a case-by-case basis, depending on the structure of the analyte. Basically it depends on the structure of your analyte and of its metabolites, and of their known or expected stability in whole blood. Which is difficult both to predict, and to demonstrate. Meaning that keeping the tubes in an ice bath, and centrifugating them ASAP after collection, would be a useful precaution. Regards Ohlbe — Regards Ohlbe |
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Helmut ★★★   Vienna, Austria, 2012-06-12 21:23 (4734 d 10:23 ago) @ konkous Posting: # 8717 Views: 5,262 |
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Γεια σου Κωνσταντίνο Agree with what ElMaestro and Ohlbe said above. It’s important to realize that only the analyst can tell the clinician how to treat samples. Some people simply ignore stability issues based on the weak excuse “we don’t have an analytical method for whole blood”. But it’s simple (use at least triplicates):
— Dif-tor heh smusma 🖖🏼 Довге життя Україна! ![[image]](https://static.bebac.at/pics/Blue_and_yellow_ribbon_UA.png) Helmut Schütz ![[image]](https://static.bebac.at/img/CC by.png) The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes |
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Ohlbe ★★★ France, 2012-06-13 20:24 (4733 d 11:23 ago) @ Helmut Posting: # 8730 Views: 5,255 |
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Dear Helmut, ❝ But it’s simple (use at least triplicates): ❝ • spike whole blood at 37 ℃* as you would do with plasma, homogenize ❝ • centrifuge one set immediately ❝ • keep the others at ambient temperature (let them cool down) and centrifuge sets after e.g., 5, 10, 15, 20, 30, 45 minutes That's indeed the easiest solution, but not always applicable. If your molecule has a high distribution or binding to the red blood cells, it may take some time after spiking to reach an equilibrium. In that case you will see a decrease of the plasma concentration over time, which will not be due to instability but to this partitioning or binding. That's one of the reasons why the EBF recommended to use a "qualified" whole blood method. However this recommendation is discussed. In the same issue of Bioanalysis journal the GCC recommended your approach, and to only go for a direct evaluation in whole blood if a partitioning or binding problem is expected or suspected. Regards Ohlbe — Regards Ohlbe |
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Helmut ★★★ Vienna, Austria, 2012-06-14 16:17 (4732 d 15:29 ago) @ Ohlbe Posting: # 8737 Views: 5,183 |
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Dear Ohlbe! ❝ That's indeed the easiest solution, but not always applicable. If your molecule has a high distribution or binding to the red blood cells, it may take some time after spiking to reach an equilibrium. In that case you will see a decrease of the plasma concentration over time, which will not be due to instability but to this partitioning or binding. You are right! ❝ That's one of the reasons why the EBF recommended to use a "qualified" whole blood method. However this recommendation is discussed. In the same issue of Bioanalysis journal the GCC recommended your approach, and to only go for a direct evaluation in whole blood if a partitioning or binding problem is expected or suspected. Thanks for the article – quite interesting! The latter case will be challenging, though.  Imagine a method validated for plasma and applied to whole blood
— Dif-tor heh smusma 🖖🏼 Довге життя Україна! Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes |
Ing. Helmut Schütz