Centrifugation and Aliquoting of Blood Serum and Plasma 

Thanks. I'm glad this helped.

I'm glad it helped!

(:

@@thunder440v3 :)

I'm glad it helped.

You are welcome!

I'm glad it helped.

I'm glad it helped!

I'm glad it helped.

You are welcome.

Thank you. This is a known error in this video. I attempted to provide some information on this error in the description.

@@VivoPhys ...and i made two corrections on my comment, huh now it looks much better...

Thus, in plasma, when blood is collected for anticoagulant, there will be no platelets under appropriate centrifugation conditions, but there will still be inactive coagulation factors (proteins of the blood coagulation system, including fibrinogen). When collected on a clot, not only platelets clot, but also fibrin from fibrinogen after activation of the coagulation factors, so the serum contains neither platelets nor fibrinogen.

I'm happy it helped.

I'm glad it was helpful.

You are welcome!

You are welcome.

You are welcome!

You are welcome.

It's hard to put a firm number on it. Itty depends on the tests you plan on doing. Some things degrade faster than others.

I'm glad it helped.

I hope this helped.

@@VivoPhys I'd did! :]

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I'm glad it helped.

It all depends on the type of analysis. Sometimes it is ok to sample the serum directly from the tube, but you must be careful not to disrupt the cells or mix the sample. I'm guessing most analyses should be complete in an hour or less.

I'd say 5 to 10 minutes but it depends on sample type and intended analysis.

I hope this is helpful.

I'm not real sure. Probably a few thousand dollars.

@@VivoPhys Do you know what model it is?

I don't. Unfortunately I've been working from home due to COVID-19 so I'm unable to check to. Sorry. Any centrifuge with cooling capabilities probably works as well as this one. This was but used. So it is not a new model.

They can be purchased from medical supply companies. I'm not sure if the color coding is universal across companies.

Whole blood is used for some tests like hemoglobin and hematocrit.

Thank you!

You are welcome.

You need to pipette it. Otherwise you will likely mix you sample. This is especially true in a anti coagulant tube.

Heparin tubes are common, but you should check with whatever company performs your analysis or makes your analysis kits/machine.

Sorry I'm not sure I'm familiar with what you are talking about. Hopefully someone else can respond to help you.

Thanks for responding so quickly. I'm a Phlebotomy student and we are learning about the different tubes & additives. I need to draw a diagram of a Li Heparin Plasma Separator Tube (PST) with gel added to form a barrier after being spun down. I searched google images & think I found what I'm looking for.

Hi Kimi O. I do not know how much red blood cell in your plasma sample is "too much", but I would assume you are correct that it does not take much. The reason is the contents of the cytosol in the red blood cell is different from the contents of the plasma/serum. So if you get red blood cells into your plasma/serum sample they will likely hemolyze (burst) releasing their content. This can dramatically change the level of some things your attempting to measure like plasma/serum potassium. So you will get an inaccurate result.

@@VivoPhys thanks so much!!

I'm not familiar with the setting you are referring to.

I have done it when necessary, but you run the risk of hemolysis which can alter the values. I would add a comment to the data in case the values are odd.

I'm glad it helped.

I'm happy it helped.

Counter weighting is important.

Hi Sɪᴢᴜᴋᴀ Mɪɴᴀᴍᴏᴛᴏ. White blood cells are part of the "buffy coat" that separates the red blood cells from the plasma or serum.

Sorry but I'm not sure. It is best to consult the manual for the test you are running. Hopefully someone else will respond who is familiar with what you are doing.

I'm glad it helped.

If you want the blood to separate then you must centrifuge it. If you want whole blood like when doing hemoglobin assessments then you should not centrifuge it.

That what the lab will be testing

It depends on what you are trying to measure in the blood. Your best bet would be to contact the company that makes your assay kit or machine. However, if the blood is exposed to the air then it is also exposed to oxygen.

Hi Nɪᴋɪᴛᴀ Rɪᴄᴇ. I do not know the process of testing blood for HIV. However, typically inverting the sample is just to mix the chemicals in the tube with the blood to either promote or inhibit clotting. I cannot imagine that it would cause an error in the test. At worst, it would probably just ruin the blood so it could not be tested at all.

Is it working though?

That's what I did

what tubes did the clinic use to extract your PRP? Which ones will you use?

Sorry, I'm not familiar with the methods you're referring to. Hopefully someone else will reply with a better answer.

You centrifuged for two long or you didn’t centrifuge straight away

I hope it helped.

I do not know. Sorry.

You can but you run the risk of hemolysing the blood cells.

I have no idea. Sorry.

You would need to check the assay kit you use for those specifics.

I hope it helped.

Sorry. I am not sure. Hopefully someone else will respond. If you find this information elsewhere please respond here so others can find it.

I have no idea. Sorry.

I hope it helped.

@@VivoPhys it did thank you!

I don't know. I have not done that assessment before.

There are several tests that may require it. It would be hard to give a list. Sorry.

@@VivoPhys its okay..

I have no experience with that vein. So I do not know.

Ideally you should centrifuge the blood and remove the serum immediately. Then the isolated serum can be stored or frozen for later analysis. Keeping the blood cells in the sample you run the risk of hemolysis.

@@VivoPhys Thank you sir. How long can I hold for the blood sample to separate from the serum? Can I centrifuge the sample immediately.?

I can't say for sure but you can probably hold onto the blood for an hour maybe even 2 hours before centrifuging. If you are attempting to get serum (not plasma) then you need to wait until the blood is coagulated. That usually takes about 10-15 minutes.

I spent some time looking this up and found a lot of poor explanations out there. Hopefully someone can double check my answer to be sure it is accurate and maybe give a better one. Regardless, here is my best attempt to explain (and understand) this. The centrifuge tube holders apply a horizontal force on the tube to make it move. Because the tube is attached to a circular object that can only spin (assuming it does not break) there is a simultaneously applied force of the tube holder pushing the blood tube towards the center of the centrifuge preventing the tubes from flying away. This force is called centripetal force. Applying these two forces simultaneously causes the tubes to spin instead of moving in a straight horizontal line. Remember that inertia = mass x acceleration (in this situation the blood tube is constantly accelerating from the forces already described or it would not go in a circle). All parts of the blood have a similar acceleration because the whole tube is moving as a unit. Keep in mind the tubes of blood are spinning with the bottom of the tube facing towards the outside of the circle, and blood is liquid (plasma) with blood cells suspended throughout. The components of the blood (e.g. red blood cells) have more mass than the liquid portion (plasma). Going back to the inertia equation, the key difference between the various blood components is there mass not their acceleration. Therefore, the more mass they have the more inertia they have. So as they accelerate towards the center of the centrifuge the components of blood with more mass (more inertia) end up migrating towards the outside of the centrifuge (bottom of the tube). This is because they are more resistant to that acceleration (i.e. more inertia) than the less massive portion of the blood.

It's not crucial. Too small can cause hemolysis, but that can happen for other reasons too. I typically go between 18-22g depending on the size of there veins.

I'm glad it helped!

@@VivoPhys so nice of you sir.. Thanks for replying to me... 😊😊

Please define PRP.

PLATELET RICH PLASMA

I'm not sure. Hopefully someone else will respond. Sorry.

To get PLASMA, the tube must have an anticoagulant. If it does not have an anticoagulant, it is called SERUM.

Good point! Just goes to prove you learn something new every day. Thanks for the comment.

I added an errors section to the description where I list this with some sources. Thanks again!

It depends on what you are trying to measure in the sample and how sensitive it is to hemolysis. Each time you spin the sample you run the risk if bursting the blood cells.

Hi Magnfiyer Lmoro. It depends on what analysis you are attempting to run. Somewhere in the range of about 10 minutes typically, but it varies.

According to BD's website: "BD Vacutainer Plus plastic serum and SST tubes are coated with silicone and micronized silica particles to accelerate clotting. A silicone coating reduces adherence of red cells to tube walls. The silica coating can sometimes cause the inner tube wall to appear cloudy and/or filmy. This cloudy appearance does not make the tubes unacceptable for use." www.bd.com/en-us/offerings/capabilities/specimen-collection/blood-collection/venous-collection/bd-vacutainer-blood-collection-tubes/vacutainer-blood-collection-tube-faq/general-tubes-faq How exactly the micronized silica particles cause platelet aggregation and clotting is unclear to me but I found this: "It was reported that SiNPs penetrated the platelet plasma membrane and stimulated a rapid and prolonged NO release, IIbIIIa activation, and finally the platelet aggregation [25]." www.hindawi.com/journals/bri/2016/2959414/

I don't know the specifics to why different protocols use different speeds and durations exactly. You would need to investigate each protocol to determine why.

I mean velocity is important to the sample. If the speed increases or the speed decreases, this will negatively affect the sample. Is this correct?

This answer is from second hand knowledge so it may not be as accurate as it could be. Too fast and it could damage delicate structures like cells or separate things that you don't want to separate in the analysis you are currently doing. Too slow and it's not going to adequately separate the things you want to separate. What is considered too fast or too slow is analysis specific.

Thank you .that was your kindness 🌼

While that improves the safety, that is not a universal requirement.

Vivo Phys - Evan Matthews it is a requirement if you are following NIH, CDC, and OSHA guidelines.

If possible please provide a link to the guidelines you are referring to help all viewing this video be as informed as possible. To my knowledge, you are not required to have the centrifuge inside a biosafety cabinet during operation. However, this would be helpful. You are required to take precautions to prevent the spread of potential aerosols. If a centrifuge you are using is not well sealed it should be operated in a micro-environment like a biosafety cabinet. Most large centrifuges will not fit in these cabinets. The OSHA website gives the advice to leave the tube in the centrifuge for 10 minutes after spinning to give time for the potential aerosols to settle. You can find this at the link below. www.osha.gov/Publications/laboratory/OSHAquickfacts-lab-safety-centrifuges.pdf

Vivo Phys - Evan Matthews CDC BMBL: cdc.gov/labs/bmbl.html

@@elizabethporretta4483 Thank you for contributing to this conversation with this link. Within that site I found the PDF below that contains all the CDC guidelines. I have not been able to find anywhere in there that it specifically says blood with unknown infectious status needs to be centrifuged in a biosafety cabinent. Can you please see if you can do a search and reply with the page number? I'm always looking to improve my knowledge and operating procedures. So thank you for your efforts! www.cdc.gov/labs/pdf/CDC-BiosafetyMicrobiologicalBiomedicalLaboratories-2009-P.PDF

Your best bet is to use a fine point marker to write on the tubes so you know which is which.

To be clear. I am not a medical doctor. I have a PhD. Drawing blood and processing it can be done without a degree.

I am in nursing school and so far what I have learned is the nurse takes the blood, sends it to the lab and the lab tech does this. hope that answers ur question! lol i'm like 4 months late responding but just came across this video!

Often, neither doctors or nurses. The person who draws the blood varies between hospitals, and may vary from time to time in the same facility. The blood may be drawn by a certified medical laboratory scientist (MLS, bachelor's degree laboratory professional), a certified medical laboratory technician (MLT, associate's degree medical professional), or a phlebotomist (may be on the job trained, or a certified graduate of a phlebotomy program), or a nurse or physician (limited training in blood collection in nursing or medical school). Centrifuging and separating the blood is done in the laboratory, and again, depending on the facility, the person who does this may be an on the job trained laboratory assistant, may be an MLS or MLT, or a combination. Who does the TESTING of that sample? That will be a college-educated laboratory professional. It may be a certified medical laboratory technician (MLT) with an associate's degree, or a certified Medical Laboratory Scientist (MLS) with a bachelor's degree.

I'm glad it helped.

I liked it video very much

Keep posting

@@Yogaabhyash Thank you. I try my best to create and upload content when my day job is not demanding. With the current COVID-19 pandemic I am much busier than normal. I will start posting again as soon as I can. Stay safe!