Educators face the issue of limited resources, so Medical Laboratory Science schools are able to illustrate only basic coagulation concepts in the student lab. George monitors the MLS Educators’ list, CLSEDUC, and came across this helpful suggestion from Program Director Michele Harms, MS, MLS. Michele gave me permission to reproduce the message and her laboratory procedure, attached at the end.
We do tilt-tubes in our student lab. The students actually enjoy these labs since they can see the clot form and love it when their tests in duplicate match perfectly. The student eye is not as fast as an instrument, so students may have slightly longer results, by a second or so.
For the mixing study lab, I use factor deficient plasmas that I re-label as patients. You could just use a heparinized sample, just don’t allow the 1:1 mix to incubate or it may act like an inhibitor. After performing a prothrombin time (PT) and activated partial thromboplastin time (PTT, APTT) with controls, they mix the patient specimen with pooled platelet poor plasma (PPPP). They run the PPPP in a 1:1 mix of patient and they see the correction, proving a factor deficiency.
Since I use factor-deficient plasmas, my students then utilize the labeled factor-deficient plasmas as reagents to mix 1:1 with their patient to discover which factor is missing. No correction = found the deficient factor. If you use a heparinized sample, you won’t have just one factor missing, so you won’t be able to do this part of the lab. The students utilize their coag pathway to determine if the missing factor is from the intrinsic, extrinsic or common pathway so they aren’t blindly picking reagents.
I also give them a brief case study on their patient, so they also know if it is a clotting or bleeding issue and the possible inheritance of the deficiency. Some of the case studies give away the disorder (clotting for XII), but they still have to prove it and they love when they get it right.
Here is Michele’s procedure: M Harms Mix Studies
Thanks to Michele Drejka for providing this interesting glim
Thanks to Michele Drejka for providing this interesting glimpse into the time-honored “factor substitution assay” profile. This is an interesting virtual exercise for students. I’ve from time to time conflated “factor substitution assays” with “mixing studies.” The factor substitution assay mixed aged serum and BaSO4-adsorbed reagents with patient plasma for detection of single factor deficiencies, as Michele has illustrated, whereas mixing studies use a normal control plasma, the standard methodology Michele and all of us use routinely. The factor substitution assays were used when we lacked rapid and reproducible clot-based and chromogenic factor assays, and we no longer test for factor substitution knowledge on certification exams.
This is a little long but I hope you find it useful: After t
This is a little long but I hope you find it useful: After the mixing study lesson I give my students the following “COAG EXERCISE” brain teaser which I composed many years ago. I tell them that with a few reagents and equipment they can use the tilt tube to figure out the exact factor deficiency of their sample. Picture yourself on the banks of the warm Amazon. There are seven mixing scenarios below each with clues to consider. Only one question will have two possible answers, the other six will be one factor deficiency.
COAG EXERCISE
BACKGROUND INFO-
AGED SERUM OR ABSORBED PLASMA* mixed 1:1 with abnormal patient plasma can provide an inexpensive although not quantitative method for identifying factor deficiencies. Using the information given below identify the missing factor(s). For more info on aged serum and absorbed plasma see Barbara Brown, 1988, Fifth ed., Hematology, p 267
AGED SERUM(AS): Rich in Factors VII, IX, X, XI, XII. Deficient in II, V, VIII.
ABSORBED PLASMA(AP): Rich in factors V, VIII, XI,XII. Deficient in II, VII, IX,X. (PIVKA)
(Both AS & AP have XI, XII, neither has II.)
Normal PT/PTT = 11-14 sec/25-36 sec.
1. Given: patient PT/PTT= 20/80sec
Aged serum mixed with patient, PT/PTT=14/33 sec
Absorbed plasma with patient, PT/PTT=22/84 sec
Which factor is missing?
2. Given: patient PT/PTT=29/42 sec
Aged serum mixed with patient, PT/PTT=30/43 sec
Absorbed plasma with patient, PT/PTT=31/41 sec
Which factor is missing?
3. Given: patient PT/PTT=13/100 sec
Aged serum mixed with patient, PT/PTT=13/105 sec
Absorbed plasma with patient, PT/PTT=13.5/28 sec
Which factor is missing?
4. Given: patient PT/PTT= 12/120 sec
Aged serum mixed with patient, PT/PTT= 12/32 sec
Absorbed plasma with patient, PT/PTT= 13/36 sec
Which factor is missing?
5. Given: patient PT/PTT=20/50 sec
Aged serum mixed with patient, PT/PTT=21/65 sec
Absorbed plasma with patient, PT/PTT=12/33 sec
Which factor is missing?
6. Given: patient PT/PTT= 13/104 sec
Aged serum mixed with patient, PT/PTT= 12/33 sec
Absorbed plasma with patient, PT/PTT=14/110 sec
Which factor is missing?
7. Given: patient PT/PTT= 40/29 sec
Aged serum mixed with patient, PT/PTT= 12/30 sec
Absorbed plasma with patient, PT/PTT= 43/31 sec
Which factor is missing?
ANSWERS: 1. X; 2. II; 3. VIII; 4. XI or XII;
5. V; 6. IX; 7. VII.
One picture is worth a thousand words! What a great fun lab
One picture is worth a thousand words! What a great fun lab that must be. I agree that a tilt-tube clot is very exciting to see. I have pathology residents rotate through my lab, and one of my opening demos is to let them do a thrombin time the old way, timing the clot and eventually seeing the fibrin strands. At first I thought this would be too elementary for the young doctors, but they all seem to appreciate seeing what goes on behind the ‘smokey doors’ of the instrument.