shujak1 ☆ 2010-06-04 12:17 (5066 d 20:50 ago) Posting: # 5434 Views: 8,360 |
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Hello All, I need to know that is it necessary/recommended to do repeat sample analysis & incurred sample reanalysis from replicate samples? Ragrads, Shuja |
Ohlbe ★★★ France, 2010-06-05 02:23 (5066 d 06:43 ago) @ shujak1 Posting: # 5437 Views: 7,182 |
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Dear Suja, No official recommendation that I know of. The problem with using the same replicate as for the initial analysis is the extra freeze/thaw cycle. Even if during your validation you have shown that your freeze/thaw stability is within acceptable limits it does not mean that there is no effect at all. By using the duplicate sample you eliminate this possible source of variability. Regards Ohlbe — Regards Ohlbe |
shujak1 ☆ 2010-06-08 08:05 (5063 d 01:01 ago) @ Ohlbe Posting: # 5462 Views: 6,943 |
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Dear Ohlbe, Thanks for the reply. Still I need more clarity on your point, ok i agree freeze/thaw effect is there and also for repeat analysis replicate aliquots can be taken,but what about incurred sample reanalyis will it be ok to do analysis from replicate samples as that(replicate)would be almost equivalent to new samples. And if we are already doing 5 freeze/thaw stability then can this effect be eliminated. Hope I am clear on my part, I need to know your point of view on this. Thanks, Shuja |
Dr_Dan ★★ Germany, 2010-06-08 14:08 (5062 d 18:58 ago) @ shujak1 Posting: # 5467 Views: 6,922 |
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Dear Shuja In some cases Ohlbe's suggestion to perform the incurred sample reanalyis from replicate samples could be right. I made the bad experience that the incurred sample reanalyis from the same samples failed but the incurred sample reanalyis from replicate samples passed. It was impossible to find out the reason for this observation but the difference was significant. We suspected additional metabolizing steps due to freeze and thaw. This was not observed during method validation. Kind regards Dan — Kind regards and have a nice day Dr_Dan |
Ohlbe ★★★ France, 2010-06-08 20:35 (5062 d 12:31 ago) @ Dr_Dan Posting: # 5479 Views: 6,975 |
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Dear Shuja and Dan, ❝ We suspected additional metabolizing steps due to freeze and thaw. This was not observed during method validation. Very interesting point. It may make me change my recommendation:
Regards Ohlbe — Regards Ohlbe |
shujak1 ☆ 2010-06-09 08:06 (5062 d 01:01 ago) @ Ohlbe Posting: # 5483 Views: 6,841 |
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Dear Ohlbe & Dan, This discussion is leading to another interesting point that whether replicate aliquots are required for such experiments (repeat/incurred sample reanalysis) or whether to have a backup of the samples these replicate aliquots are prepared and stored at same temperature in different deep freeze apart from where original aliquots are stored, may be for proper control of samples (we call them analytical and clinical aliquots respectively). As one of the European auditor asked to have these separate aliquots for backup purpose. What's your view on this. Regards, Shuja |
Dr_Dan ★★ Germany, 2010-06-09 13:30 (5061 d 19:36 ago) @ shujak1 Posting: # 5485 Views: 7,393 |
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Dear Shuja I also made the experience that auditors asked to have separate aliquots for backup purpose. They want to be on the safe side but they are wrong: Once the study report is finalized you do not need the samples anymore. If the study passed everything is fine, if the study failed you would not be allowed to do re-analysis. If the study is subject of an inspection and you still have the samples what would be the advantage? At that time the stability would have been exceeded. So my suggestion is to keep the aliquots in two different places (freezers) but under the same conditions in the clinic and later in the lab. Send the first aliquot to the lab and after it arrived safely you send the second. Keep in mind that the second aliquot is usually regarded as a back up in case of damage to the first. Hope this helps Dan — Kind regards and have a nice day Dr_Dan |
shujak1 ☆ 2010-06-11 08:19 (5060 d 00:47 ago) (edited by shujak1 on 2010-06-11 10:51) @ Dr_Dan Posting: # 5498 Views: 6,851 |
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Hello All, I think we are heading towards some open ended questions with still no conclusion, mean while I got another prospective of this that it is generally asked that the bulk spiked standards/samples shall be stored along with the study samples to have/mimic same conditions as that of the study samples so my concern is that if these 2 are to be stored in same location under same temperature conditions then is it ok to keep replicate/duplicate aliquots of study samples at different location under same temperature conditions. And Yes I am still waiting for Helmut views on this whole topic ! Regards, Shuja |
Helmut ★★★ Vienna, Austria, 2010-06-11 17:11 (5059 d 15:55 ago) @ shujak1 Posting: # 5504 Views: 7,100 |
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Dear Shuja & all, ❝ I think we are heading towards some open ended questions with still no conclusion, […] Well – maybe there is none?! ❝ And Yes I am still waiting for Helmut views on this whole topic ! I changed my name above from big to small because I left the lab many years ago. Some thoughts: Bulk vs. freshly prepared calibrators/QCs. No specific recommendation in the FDA’s guidance, the Crystal City III conference report, and the EMA’s draft GL. Freshly prepared calibration curves are only mentioned in the respective sections dealing with stability testing in method validation. But: in my experience inspectors prefer freshly prepared calibration curves over bulk samples (a large volume is spiked, split into vials and frozen together with QCs and study samples). As Ohlbe once mentioned: ❝ Though inspectors are paranoiacs and won't trust anything unless it is written, an inspection remains a confidence-building process. We used bulk calibrators for many years for the following reason: Spiking small plasma volumes (with a very small volume of analyte solution) is inherently variable. Many CROs even prefer to prepare standards (and QCs) gravimetrical instead of volumetrical. Now for a nasty example: You have validated storage at –20 ℃ and store bulk calibrators and QCs together with study samples. After the last analyst left the lab (and checked the temperature of the freezer) you have a power outage. Temperature rises to –10 ℃ and returns to normal during the night. The analyte starts to degrade at >–15 ℃. If you have an online-warning system connected to your freezer, this must be battery-powered! Let’s assume you don’t have that. In the morning the analyst checks the temperature of the freezer – everything is fine. Responses in further batches will be lower than in previous ones. You will see lower CC slopes, but runs are not rejected because QCs are affected as well. If you use freshly prepared CCs instead, runs will be rejected (degraded QCs will fail). On the other hand bulk CCs are less variable and in BE – where we are interested in intra-individual comparisons – the ratio of test/reference will not be affected anyhow. At the EUFEPS/EPF-workshop last April Kamal Midha and myself made an argument in this direction, but it was obvious that regulators didn’t like that at all. Back-up sample vs. thawed original. I would follow Ohlbe’s recommendations. Storage of samples after the end of the study. I partly disagree with Dan’s post. In my CRO we followed this procedure:
— 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 |
tusshar_adi ● 2010-06-24 20:55 (5046 d 12:11 ago) (edited by Ohlbe on 2010-06-24 23:16) @ shujak1 Posting: # 5559 Views: 6,717 |
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It is not necessary but in some cases where stability or immediate conversion or interconversion of the analyte does make it necessary to use the repeat sample analysis and incurred sample reanalysis from replicate samples. Edit: Full quote removed. Please delete anything from the text of the original poster which is not necessary in understanding your answer; see also this post! [Ohlbe] |
Jagdish Bairagi ☆ 2010-07-27 18:02 (5013 d 15:04 ago) @ shujak1 Posting: # 5691 Views: 6,566 |
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dear suja Repeat analysis should be done from replicate samples only. |
Helmut ★★★ Vienna, Austria, 2010-07-27 18:16 (5013 d 14:51 ago) @ Jagdish Bairagi Posting: # 5692 Views: 6,476 |
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Dear Jagdish! ❝ Repeat analysis should be done from replicate samples only.. I don't understand your statement. Can you please give a clarification? — 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 |