pha/th test samples

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afhm

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How long can a sample be stored before you send it in?  Would I need to keep them refrigerated?  I have animals that need testing in several different pastures and can't get them all penned to take the samples at the same time.  It might take a couple of weeks or so.
 

red

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Afhm, I guess I would take a cooler w/ me while you were collecting the samples. I think you need to send them in ASAP after drawn. I know they recomended not collecting blood before a long weekend. Place an icepack in the box also when you send them in during the summer. I guess, I send what I collect & then send the others in as you collect them.
I'm sure if DL or Barrel Racer has other ideas, they'll let you know.
Good luck, we're drawing blood tomorrow! I'll cross my fingers for you if you do the same for me!

Red
 

DL

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Although in theory you could wait until you had all samples collected, what happens with storage (in the refrigerator) is that over time the sample degrades and then they may not be able to extract DNA which means you won't get any results which means you would have to draw them again.

I would send as I draw them, also would put in cooler (not directly on ice) as you draw them (esp if it is hot) and then refrigerate. I don't know where you are but I send my on Mon or Tues UPS ground (Michigan to Illinois) takes 2 days costs 10 bucks. In this weather you don't want them sitting around getting hot, so pack with a couple of frozen cold packs and send early in the week - if you are far away or ground takes more than 2 days I would 2nd day them...

hope that helps - let me know if you need more info (ps if you want both TH and PHA you need to indicate that)...dl
 

Jill

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I've been looking at this animal of Reds thinking she was all positive and that was a little red hen.  I just now read the caption, didn't realize that was a turkey. (lol)
 

red

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got through a little "Red" humor in there everyonce in a while Jill!
That was one of my favorite quotes when I worked!

Red
 

xxcc

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DL, correct me if i'm wrong, and Dr. Beever may not like this idea, but by rights, shouldn't one be able to ear notch an animal and place it in a sample container with a liquid like Formalin, then once all are collected, send them in? Similar to ear notching for BVD, etc.?  I would think that wouldn't degrade to much and they'd be able to extract DNA from them?
 

DL

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Hey xxcc - On TV they can extract DNA from a molecule of spit left on a door nob that is 100 years old!  In theory this is possible, however the PHA & TH tests have been optimized to use whole unclotted blood (extract DNA from white cells) or semen.  I think it would add a whole layer of complexity to do it with formalized ear notches. When the defective gene was being identified DNA was extracted from PHA calves ears (not in formalin) - I think the tissue adds additional steps. Like parental verification - some labs use blood some use hair roots. That is my understanding anyhow..When you are ear notching for BVD you are either doing immunohistochemistry (basically you add an immune probe that binds the virus if the virus is in the tissue and then lights up - each slide has to be examined under the microscope); many labs have started doing ELISAs on ear notches - which is basically an antibody-antigen reaction done in plates so a bunch can be done at a time - the preferred sample here is ear notch not frozen but refrigerated; can be frozen once, but freeze-thaw cycles degrade the virus.....probably way more than you wanted to know - so the bottom line here is blood (purple top tube) or semen - to the lab in a timely fashion.... .
 

red

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Barrel Racer- sent in 6 samples. Expressed mail them.

Red
Red Maple Farm
 

Barrel Racer

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Okay...blood storage -  the important thing is that the blood not freeze or get too hot.  The DNA in blood comes from the white blood cells, when blood freezes or gets really hot it lyses these cells which means no DNA.  In theory, as long as you keep the blood in the fridge it SHOULD be fine (I say should because sometimes they just don't work, nothing is absolute).  I have gotten blood tubes that have been out in room temp for a week to work as well as tubes that have been in the cold room for a year to work for certain things (have not done that on TH/PHA test though).  Dr. B says to send them in in a timely fashion because if you stick it on your dashboard of your truck and drive around with it there for a week, it's probably not going to work.  Another important thing is for the blood to be unclotted, blood clots cause a lot of heme in the DNA sample and the enzymes for the test do not work well in the presence of a lot of heme (heparin also kills the reaction completely so do not add heparin). 

On the tissue samples - we have to isolate the tissue samples a completely different method and honestly it's not cost or time effective for high throughput samples, meaning the cost for you as a submitter will have to go up substantially.  The reagents required add quite a bit of additional cost as well as the cost of labor involved in working with tissues.  It takes a lot more time to isolate DNA from tissue or hair than blood or semen.  The TH/PHA tests are extremely finicky as well so any contaminents (hair samples are notorious for having contaminents) can cause the test to fail.  In my experience the tissue samples do not yield good high quality DNA, the ears that we used to do all of the research were in general poor quality and some of the DNA just would not work at all.  I had to use a  DNA amplification kit (which is what the crime labs use on everything) on what DNA I had and that seemed to work great, but that kit costs about $50 per sample just for the amplification kit.  A DNA test is completely different than a BVD test (DL explained pretty well). 
 

knabe

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as someone who works in a genome center, and also with plants in the past, it is convenient to require all samples be the same.  this way you have a smaller set of problems to troubleshoot when things dont' work.  also, when results are much more predictable, scorable, all reducing the time to return results.  this by the way is always a problem in DNA type situations.  at some point, every sample has to be standardized, and the earlier the better, particularly if it is before the center receives it.  this way, you only need one pipeline, one set of protocols, less legacy knowledge, greater ease of transfer of knowledge, overall lower costs.  also, the blood sample allows one to forward some of that on for more bloodwork for genoypting for ET requirements.  one could also use the sample for a future test like monkey mouth, etc.

when one must use different types of samples, it rarely is streamlinable, so it is much easier from the "bigger" picture, to force (force?) the collector to conform to a protocol that is easiest for the lab to process cheaply, quickly, uniformly.

monkey mouth, when for monkey mouth????????????????
 

red

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never have seen a picture of a calf w/ monkey mouth. anyone got a picture of one?

Red
 

Barrel Racer

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knabe said:
monkey mouth, when for monkey mouth????????????????

So many defects and only one of me  (lol)  I'm gearing up to start epilepsy in Herefords next.  We've got quite a few defects on the list, monkey mouth being one of them white heifer disease, sp, and a few others too.  The problem lies in the search for samples and manpower.  I've got some samples for monkey mouth, but not near enough to start mapping with them.  It took about 6-7 months just to get samples collected for TH and PHA, so probably won't be any time soon for mm  :( (unless someone else is working on it, but to my knowledge no one is). 
 

red

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Epilepsy in Herefords? That's one I'd be really interested in since my niece & I both have problems w/ seizures. Can you send me more information?

Red
 

afhm

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Barrel Racer so can I take samples and refridgerate them for about a week before I send them in and still be OK?  I 've got 10 head to take samples on, but they are in 4 different pastures several miles apart and of course only 2 of them have a chute to restrain them in.  I can get 4 of them tommorow, 1 of them at anytime, and the others may take me a few days to get to them.  Iwant to send them all in at the same timeso I can get the results at the same time and save on the postage.  Thanks.
 

Barrel Racer

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afhm said:
Barrel Racer so can I take samples and refridgerate them for about a week before I send them in and still be OK?  I 've got 10 head to take samples on, but they are in 4 different pastures several miles apart and of course only 2 of them have a chute to restrain them in.  I can get 4 of them tommorow, 1 of them at anytime, and the others may take me a few days to get to them.  Iwant to send them all in at the same timeso I can get the results at the same time and save on the postage.  Thanks.

You should be fine, just keep them in the fridge and if you think about it take a cooler with an ice pack in it with you when you collect them, depending on the weather.  I'd stick an ice pack in when you ship them, it's been upper 80's here and supposed to be over 90.  I think Dr. B uses the rule 90 and above to stick in an ice pack when shipping. 
 
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