Major Research Problem!!!

[longhorn]

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I am workin at an NIH lab this summer and have come across a problem in the last week. I am not going to go into too much detail but I basically found out the cell line our lab is using has mutated and has lost expression of a major oncoprotein. I told my postdoc and we discussed it with others who work with the cells. One of the guys was really emotional about it and ran to validate my results. The other, didn't seem to care, told me he had been noticing some strange protein levels in these cells. He then procedded to tell another senior scientist the next day that everything was fine. I then asked him for a lysate of his cells to test since he wasn't going to, and he rufused. Finnally, he gave me some cells which i had to grow, harvest, run a protein determination on, and a western. Well, I did some investigating of his cells and some older feeze downs, one in particular dating back to 4/18/01 and have found none of the cells express this protein anymore. The big problem with this is this cell line was made by my PI and sold to a company so anyone can use them. Now that means that any published papers by our lab could be refuted by another lab who has the normal cells. Another problem is the wasted work that has gone on since at LEAST 4/18/01. And this is government money too. so we all foot the bill. I have talked to my postdoc about telling the PI about this when I present my data for the summer(this week). He at first told me not to, saying you have to be a team player and how timing is everything. I totally understand this but I think this serious of an issue needs to be brought to the boss. Also, it doesnt seem like my lab is working as a team on this issue and I think some authority is needed here. If it was something minor I wouldn't mind "hiding" my results. This decision to tell or not is really weighing heavily on me. I do not want to start my career off on this kind of note and I definetly wish I had not found what I did.

Another twist to this is my PI is a real hot shot(Lets just say he is Editor in CHief of a really really big name journal and one of the top doctors in his field). IF I keep this from him it means any research I did with this cell line is not accurate and I can not in good conscience present it to him. I will not put my name on any data which I know is a misrepresentaion. This makes my research for the summer half as good as it should be(I was using another cell line as well which is still good) and also eliminates most of my chances for a publication. I was planning on asking my PI for a rec. for medical school but I know if I completely omit this cell line my research doesn't look so good and either will the rec. I am also fully aware of how good my rec will be for finding this out and telling him.

In summary I was planning on telling everyone in my lab my results and my intentions before I tell the PI. I think that is a fair solution to this problem so everyone has a chance to be prepared for any repurcussions. If i do not tell I get screwed and I guess my lab has more time to find out what happened before they tell the PI. If I tell I will look good and at least the problem will be handled properly. I hope I am making the right choice and not just the one which will benefit me the most. What do you guys think and any ideas on how to bring this up????? THANKS

 

[Bikini Princess]

Whoa..I would definitely be diplomatic about this. Retain your credibility, and remain somewhat reserved with own your feelings. (i assume you are, but it's probably hard). Don't come across as taking one side or the other, simply present the data to your PI, with a modest conclusion.

You've been doing controls with all these antibodies, I hope? ie. cell lysates from other cell lines which do express the proteins, in the same westerns?

What is the evidence that this oncoprotein has ever been expressed in your line? depending on your antibody (mono or poly clonal?) the mutation may be epitopic, preventing antibody binding, this might not alter the protein function too badly, so things might not be as bad as you think..

 

[longhorn]

I have tried this with many different lysates even using different lysing procedures. We know both cell lines had what I would call blazin' amounts of this protein in 2000. I have ran positive and negative controls on both cell lines together in the same gel. Trust me the last thing I want to do is look like an idiot here. Your antibody reasoning is one I have thought of and is the only way I could be wrong. But I do know this same antibody picked up both cell lines in 2000. I forgot to add this, there is some of this protein in the mutated cell line but it is very very weak. I have to expose for 10 minutes using ECL+ just to get the hint of a band. I have also run B-tubulin Western to make sure there is protein in these mutated cell lanes and there is equal amounts in both cell lines.

 

[longhorn]

and i am using a very popular mouse monoclonal antibody

 

[Bikini Princess]

I'm guessing, is it Myc?

It seems odd that the line would do this knock-out mutation by itself..usually these are pretty redundantly encoded proteins anyway, right? Have you considered if post-translational modifications might be affecting the epitopic binding of ab, since it's monoclonal? (ie do you know where the epotopic region is? maybe it's prone to cleavage, so might be as simple as needing to replace your protease inhibitors).

I suspect that complete mutational deletion of protein expression is a "zebra", but of course zebras do happen.

 

[longhorn]

Good question. I will look into that. And its not Myc.

 

[Bikini Princess]

Is it p53?
ok i'm getting off the computer to go work out now. too many essays, too much sdn.

 

[exigente chica]

That's a really hard decisison to make. Last year, before starting my research, I had to take a course called "Integrity in Science"..we watched this movie and it showed how scientists cheated or did sloppy work and still published datat. It also showed how grad students would tell on their PI and the outcome was very nice, they were discredited and had to find another line of work.
It's hard..umm. I don't know what to tell you...mann..it's hard..and your working at NIh, ooh😕

 

[Vader]

Sounds like a very tricky situation. I would be very sure before bringing something like that up to the PI. If you feel you need to do so, then bring it up in a non-judgemental, objective manner. It is important to do what is ethical in this situation, so as not to hurt your own career down the road. The long-term reprecussions of not disclosing this information could very well be ultimately more damaging than being diplomatically and politely forthcoming.

Good luck... hope it all works out.

 

[none]

I think you need to tell the PI, but you need to do in a way that does not go over the head of the postdoc you work under. You need to be clearer to your postdoc that this a major concern of yours. Explain in detail what your concerns are. You seem quite frantic in your post... The publication, while definitely beneficial, is most definitely not required for MSTP admissions (I can personally attest to this!) I think your letter of rec could still be quite good even if one line was a complete failure. Letters of rec don't have to be all about results. You'll have plenty of time to discuss results in your interviews. Really good letters of rec attest to things like character, personability, work habits, career interests, etc...things that aren't really all that science related. Honestly, it sounds like you have A LOT of research experience behind you and MSTP admissions aren't going to be a problem no matter how this works out.

 

[kutastha]

I posted in your other thread, but I'm curious why you're not interested in why p53 expression is lost in these cell lines. Depending on your lines, it sounds like a fairly decent project in itself, and would help in answering a lot of your concerns that you've brought up.

 

[longhorn]

yea no kidding. We joke around that this is exactly what we are looking for, loss of p53 expression. But I need to deal with this political bs before I can jump on the research.
BTW I am not MD/PhD. ITs funny how experiences in your life formatively mold your popinion.

 

[kutastha]

Well, the politics of your lab sounds like the fox is watching the henhouse. After I posted last night, I was thinking of the multitude of questions your results provide - did your control lysate on your Western (or ELISA?) show p53 expression? What kind of cells are you using? What passage #? Were they near crisis? Have you done an RT-PCR to test for the message? You said a mutation was responsible for KOing your p53 - what mutation?

These questions may not be your responsibility and the person you answer to should either (a) know them, or (b) ask them of you as well. If it's neither, you may need to set up an appointment with your PI to discuss your results and not dabble in the politics of the lab.

 

[longhorn]

SAME POST AS IN MD FORUM

Ok, so I finnally told my PI today. This guy is a class act. You could tell he was furious but he still kept his kool in front of me, only slipping up a few times with some obviously pointed comments to those responsible. I basically did what you guys said, came in there humble, and with the belief the data are correct. He was happy with me, but focused more on what was going to be done. I definetly have this rec sewed up though.

Looking back I am glad I played things the way I did. I wish the correct answer would have struck me more as the correct answer, but I guess thats why decisions are tough. I do feel bad about any repercussions from this event, but at least I know this situation is now a top priority.

BTW my postdoc told me later there was a sentiment of guilt and fear in those repsonsible for what happened. He told me in no way were they trying to hide any data, rather just trying to delay its disclosure until more evidence had been gathered. I am still shocked that a discipline as pure as science, based on logic and strict naturalistic principles, is still just as easily corrupted by the pressures of politics and bureaucracy as any other field. I guess no matter how fair and "scientific we claim to be, we still are as bad as my hometown company, ENRON.

 

[longhorn]

Oh most importantly, Thanks to all who gave input, it was greatly appreciated.

 

[Bikini Princess]

Originally posted by longhorn
SAME POST AS IN MD FORUM

Ok, so I finnally told my PI today. I basically did what you guys said, came in there humble, and with the belief the data are correct. He was happy with me, but focused more on what was going to be done. I definetly have this rec sewed up though.

sounds like things will work out ok then. 🙂