|WikiProject Molecular and Cell Biology||(Rated C-class, Low-importance)|
|WikiProject Viruses||(Rated C-class, Mid-importance)|
Elution using trypsin
I read that phage can be eluted with trypsin, but if you do that, and it cuts off the displayed protein, then how can you repeat the panning? Coat protein III which is usually what binds the displayed protein to the phage only occurs in one place with upto five units, and rounds of panning are often completed more than five times..? --Username132 16:19, 26 January 2006 (UTC)
The coding information for the expressed protein is contained within the phage particle. When the eluted phage are re-infected into host bacteria the displayed protein is expressed once more and displayed on the newly produced phage. Phil Scrutinator (talk) 17:41, 14 March 2008 (UTC)
KM13 is a popular helper phage that allows for elution using trypsin (although my experience suggests that elution be performed using TEA first, then trypsin treatment). Although this cuts the displayed protein, the genetic information is still contained within the phage. Once you infect bacteria with the eluted phage and amplify, the protein becomes displayed on the coat protein of the amplified phage for subsequent rounds of panning. Hope this helped. 184.108.40.206 17:47, 14 February 2006 (UTC) Warren D. Marcus
Description of phage display
I think that it is wrong to say that phage display is a method to study (protein/protein or protein/DNA) interactions. I would say that it is a method for selecting for a particular protein or peptide phenotype while maintaining the link for the genotype. Thus, it also allows for multiple rounds of selection and (possibly error-prone) replication, mimicking evolution in the lab. One straightforward means of selection is indeed to use the interaction of the protein or peptide with some other protein, or DNA. But it's also possible to select for other features. For example, you can select for protease resistance (with an affinity tag which is or is not cleaved off). This is indeed mentioned at the end of the article (protein engineering), but the wrong definition at the beginning masks that more general meaning of the term.
In that context, one interesting information is missing: Which Phage display systems can be used for which sizes, what's the largest protein to be possibly displayed? Unfortunately, I don't know the answer (actually that was the reason why I visited the entry).
I don't think there is a simple answer to this. The two predominantly used phage - T7 and filamentous M13 derivatives - differ in their size limits and it also depends on the nature of the protein and the coat protein used. Certainly both cpIII and cpVIII can display Fab heavy chain - approximately 800 bp of coding sequence. Another factor is whether the phage uses a mixture of wild type and fusion coat protein or just fusion coat protein - this can affect the infection process. The pJuFo system of Reto Crameri can potentially express whole cDNAs by using fos and jun leucine zipper fragments as intermediaries to cpIII. Phil Scrutinator (talk) 17:41, 14 March 2008 (UTC)
The article does seem to focus on filamentous phage display and accentuates the need for helper phage. This ignores methods based not on phagemids but on engineered M13 phage - the approach taken at the outset by Smith. Phil Scrutinator (talk) 17:44, 14 March 2008 (UTC)
Pieczenik patent litigation
Discussion for the material relevant to this topic. Licenses/Patents/Disclaimers: Ph.D.™ is a trademark of New England Biolabs, Inc. This product is sold for research use only and not for resale in any form. Commercial use of this product may require a license. For license information under U.S. Patent No. 5,866,363 please contact the Licensing Office, New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938.
Commercialization of sequences covered using this product may require a license from Dyax Corp. under U.S. Patent No. 5,403,484 and associated patent rights. For license information contact the Director of Corporate Development, Dyax Corp., One Kendall Square, Bldg. 600, Cambridge, MA 02139, Fax 617-225-2501. Good luck to you too. — Preceding unsigned comment added by GPieczenik (talk • contribs) 00:20, 10 May 2012 (UTC)
Pieczenik patent litigation
Latest edits removed with reasons.
Phage Display was originally invented by Prof. George Pieczenik in 1983 USPTO disclosure document 118831 [] cited as proof of date of conception in a continuation-in-part US Patent 5,866,363 originally filed in August 1985 [] [n 1] and told to Vidal de La Cruz by Prof. George Pieczenik when they met at NIH[n 2]. Vidal de La Cruz was in turn credited for Professor Pieczenik's idea by George P. Smith in his 1985 publication[n 3],
This location can only accept small DNA fragments. Therefore, this vector was not used as a display phage vector until Prof. George Pieczenik re-introduced using f1 bacteriophage at its single HinD site in gene III (which does not exist in M13) as a vector for randomly synthesized DNA fragments. This created the first phage display library.[n 4] This was done at the MRC Laboratory of Molecular Biology. George Smith only created a simple vector displaying a small DNA fragment of no consequence[n 5] until the concept of inserting randomly[n 6] generated sequences was introduced by Prof. George Pieczenik.
The invention of antibody phage display by Prof. George Pieczenik  and reduced to practice by the technical staff using Zinder's f1 bacteriophage and Prof.Pieczenik's protocols and Cadbury reagents at the MRC Laboratory of Molecular Biology led by Greg Winter and
Prof. George Pieczenik priority claims 37 USC 102(e) are validated by Nobel Laureate Bruce Merrifield [][n 7],[][n 8]. Prof. George Pieczenik has sued many infringers and will sue Wikipedia if it continues to insist on publishing fraudulant inventorship claims when its editors have no understanding of what a continuation-in-part patent application is and that the submission date refers back to the original filing. The stupidity of the Wiki editors has created alot of noise and very little accurate information. They are the dupes of the pharma corporations and paid to publish pharma propaganda.[n 9]
Updated at Staticd (talk) 10:06, 13 May 2012 (UTC) new edits by GPieczenik in question These are mostly the previous matter except these new links.[n 10] even the wiki-syntax errors are identical, indicating that GPieczenik has no intention of reading anything told to him or taking part in discussions.
Problems with edits
- the only copy of the disclosure document[r 1] I could find was on GP's personal page, the USPTO has not mentioned it nor have they archived it publicly. Also, the merrifield letter [r 2] is archived on GP's personal page. So I question their use as reliable sources for wikipedia (something a weee bit more neutral is needed) .smiths 1985 paper[r 3] was submited in 1984, accepted in april 1985 and published in june 1985. How does the patent filed first in august 1985[r 4] get priority? (taking the two RS availible : the paper and the patent) GP may be the true inventor, but I find it ridiculous that the only proof for priority are two documents on the claimant's Personal page. Some one please prove me wrong.
- . no way around this. the claim needs a reference
- most certainly not. neither GP nor V de la cruz are acknowledged or cited in Smiths 1985 paper. Or do I have the wrong paper?
-  for the (a) the publication about the library, (b) for it being the first. THe first I could find was [r 5]
- for arguments sake, 2000+ citations must be of some consequence. But claiming either way without a citation is not allowed on wikipedia as per wikipedia policy WP:OR
- "The success of the foregoing affinity- purification experiment gives hope that antibody might be used to isolate desired clones from a library of random inserts in a fusion-phage vector." The concept already mentioned in smith's paper.
- self archived and of questionable reliablity for an encyclopedic reference
- completely irrelevant. makes no mention of the issue at hand
- The burden of proof lies with the person making the claims. Please take trouble of learning about wikipedia policies when they have already been pointed out to you.
- All of them are irrelevant, both to the page in general and in the context of their insertion.
- USPTO Disclosure document 118831 (1983)
- merrifield letter
- Smith GP (1985). "Filamentous fusion phage: novel expression vectors that display cloned antigens on the virion surface". Science. 228 (4705): 1315–1317. doi:10.1126/science.4001944. PMID 4001944.
- US patent 5866363
- Parmley, S. F.; Smith, G. P. (1989). "Filamentous fusion phage cloning vectors for the study of epitopes and design of vaccines". Advances in experimental medicine and biology. 251: 215–218. doi:10.1007/978-1-4757-2046-4_21. PMID 2481962.
Here is the link that was deleted http://www2.mrc-lmb.cam.ac.uk/about-lmb/archive-and-alumni/alumni/dr-george-pieczenik/
You will notice that I am credited also for having an idea that was appropriated by 3 Nobel laureates, Francis Crick, Sydney Brenner and Aaron Klug. Remember good intention is no excuse for ignorance. Pharmaceutical companies also have good intentions..their primary one is to make money. So please don't act much abused. Best, Prof.Pieczenik Already I have enough evidence to tell my students exactly what the level of credibility is. Have you contacted George Smith and asked him if anything you claim is true. He never said he invented phage display of combinatorial libraries. He actually thanks Vidal de La Cruz in his Science paper for the suggestion. And if you ask Vidal de la Cruz he will admit my telling him about my idea and patent filing. So if really have read the references then you would see what I am saying is accurate. The file wrapper on a patent is also a legitimate reference. Just because you can't find it on the internet doesn't mean it doesn't exist. That is the problem with Wiki. Very superficial and an arrogance to maintain that superficiality. Not good. It undermines the quality of students being able to think rather than "cut and patch" from Wiki, which is now the norm. Remember people's lives are at stake in the end because this is the future of personalized medicine, which I have now created with these libraries. Most of American science has just be appropriated from others abroad. Molecular biology in particular was created and started in England at the MRC Laboratories. Everything on Wiki fails to recognize that fact. More in sorrow than in anger. — Preceding unsigned comment added by GPieczenik (talk • contribs) 23:07, 12 May 2012 (UTC)
- First, I have reverted your changes to the article. Please reach consensus here before making these edits again.
- With regard to the content you are wishing to add. Please explain how the citations you provided verify the content you provided. The citations provided do not even mention the phrase "Phage display", while the citations you removed do. I am not a biochemist (although I have now asked a couple to review this conversation as well), but I should be able to understand in basic terms how the patents Dr Pieczenik is credited with are related to the content you are adding. Please expand on that relationship here.
- Lastly, please review Wikipedia guidelines and policies that others have pointed out to you; including verifiability, reliable sources, citing sources, and conflict of interest. Also, please sign your posts using three tildes when posting to talk pages.
- Thank you, I look forward to your response here. --Tgeairn (talk) 23:25, 12 May 2012 (UTC)
- No one is trying to "arrogantly maintain superficiality". I would greatly like a better reference than something only on a personal webpage. We are going deeper (not superficial) by getting more and better references (on or offline). And by asking you to help me get them (having given both the criteria and explanations for how) where I have failed after searching in good faith, I hope I act in the truest spirit of cooperative building that is wikipedia (certainly not arrogance). If you have read my previous posts, you would notice that I mentioned de la Cruz not being cited/mentioned in the smiths 1985 paper. Again, If you have a reference please do provide it. You are making the claim, the burden of proof lies with you.
- I am not sure who deleted the alumini link or when but whatever the reason, how is it relevant to this topic? Also what does the protein synthesis paper (crick,brenner,klug) have to do with this topic? (unless you are trying to make an ad hominem argument about your greatness. that holds as much water as User:Obama )
- "Good intentions may not excuse ignorance" No one who has interacted with you has tried to excuse their ignorance. Just asked you help remedy it, repeatedly. On the other hand, having money in a bank does not excuse breaking into the vault - you have to fill the forms at the tellers (or give satisfactory references in wikipedia, even if you are the inventor).
- Calling editors pharma stooges is rude.
- "It undermines the quality of students being able to think rather than "cut and patch" from Wiki, which is now the norm." - In my experience any teacher worth their salt inspires students with their lectures sufficiently and sets cool enough assignments that (most) students gladly review literature on their own, think critically and go wayy beyond the wikipedia article on the topic (if it even exists, and it usually shouldn't with a good teacher)
- If "cut and paste" is the _norm_, in my experience, it's due to an exceptionally crappy teacher.
- Staticd (talk)
Expert assistance request
|This topic is in need of attention from an expert on the subject.
The section or sections that need attention may be noted in a message below.
The lede and Principle sections of this article contain potential original research or unverified claims. The attention of an expert in this subject is requested as I (and other editors) have been unable to verify the claims. Please use this section to discuss. Thanks, and Happy Editing! --Tgeairn (talk) 01:53, 13 May 2012 (UTC)
- I think Staticd's and Tgeairn's analysis in the previous section is not very accurate. USPTO disclosure document 118831 that would give US patent 5,866,363 priority over PMID 4001944 (Smith_1985) is a reliable source. Hence the Smith_1985 publication appears not to be the first description of phage display. Furthermore Vidal de La Cruz is not mentioned anywhere in Smith_1985. Hence I am restoring the original lead. Boghog (talk) 04:50, 13 May 2012 (UTC)
- For balance, I have also added a reference to US 5,866,363. Boghog (talk) 10:50, 13 May 2012 (UTC)
- If anyone is curious, a typed version of the hand written USPTO disclosure document 118831 may be found here (see Appendice B). This document is still not a reliable source. But even if it were, this document does not contain a clear description of phage display. The only thing that comes close is the statement "can use viral, plasmid, bacterial, parasite, any organism DNA and generate similar matrix" where the matrix is an expressed library of antigens (i.e., peptides). Hence it appears that Pieczenik had thought of the general concept of display libraries in 1983, but the initial description was very broad covering display libraries generated by essentially any organism. Furthermore there is no clear description about how this general concept could be reduced to practice using phages. So to say Pieczenik had developed the idea of phage display in 1983 is a stretch. Boghog (talk) 17:34, 13 May 2012 (UTC)
- Boghog, thank you for your clear explanation and your work to improve the article. Very much appreciated! --Tgeairn (talk) 20:24, 13 May 2012 (UTC)