(M) Lesson 8: Automation and Trends in Hemostasis Flashcards
What is the preferred method or principle of measurement in automated coagulation testing when dealing with HIL (hemolyzed, icteric, lipemic) samples?
Mechanical Method
When a clinical laboratory receives a sample that is icteric, lipemic, or contain substances that interfeere with light transmission, it is recommended that they use what method?
Mechcanical or Electrochemical Methods
According to the CLSI Guidelines
According to the College of American Pathologists, it is indicated that coagulometers using an optical detector may have problems with ____ on icteric or lipemic samples.
End-point determination
Levels of Coagulation Instrumentation Automation
- All reagents are transferred manually by the operator.
- Temperature is maintained by a water bath or heat block.
- External measurement by operator may be required
- End-point is determined visually by the operator.
- Timer is initiated and stopped by the operator.
- Examples: Tilt Tube, Wire Loop
Manual
Levels of Coagulation Instrumentation Automation
- All reagents are transferred manually by the operator.
- Instrument usually contains a device for maintaining constant 37 deg C temperature.
- Analyzer may internally monitor temperature.
- Instrument has a mechanism to initiate a timing device automatically on addition of final reagenet and mechanism for detecting clot formation and stopping the timer.
- Examples: Fibrinometer, STart 4, Cascade M and M-4. BFT-II, KC1 and KC4
Semiautomated
Levels of Coagulation Instrumentation Automation
- All reagents are automatically pipetted by the instrument.
- Specimens may or may not be automatically pipetted.
- Analyzer contains monitoring devices and an internal mechanism to maintain and monitor 37 deg C temperature throughout the testing sequence.
- Timers are initiated and clot formation is detected automatically
- Examples: ACL TOP, STA-R Evolution, STA Compact and Compact CT, Sysmex CA-series, BCS XP, CoaLAB
Automated
Instrument Malfunction Flags or Sample Quality Flags?
- Temperature Error
- Photo-optics Error
- Mechanical Movement Error
- Probe not Aspirating
Instrument Malfunction Flags
Instrument Malfunction Flags or Sample Quality Flags?
- Lipemia
- Hemolysis
- Icterus
- Abnormal Clot Formation
- No end-point detected
Sample Quality Flags
When problems in the instruments arise, the users will have to?
Call for help from the manufacturer and/or service engineers to come and repair it.
T or F: Specimen quality flags are most likely analytical errors.
F (pre-analytical errors)
Match the following.
- Falsely shortened CT due to premature activation of CF and platelets that generate Factor VIIa and thrombin. (2 answers)
- Falsely prolonged CT due to interference with light transmittance.
- Indicates liver dysfunction that may lead to prolonged CT because of inadequate clotting factor production.
- Falsely elevated CT due to instrument inability to detect an end-point.
- Indicates that the instrument was unable to detect clot formation.
A. No end-point was detected
B. Abnormal clot formations
C. Icterus (Bilirubinemia)
D. Hemolysis
E. Lipemics
F. Clotted
- F, D
- E
- C
- B
- A
If the result obtained in a photo optical principle, then ____ princirple may be used as alternative.
Mechanical
If both mechanical and optical are unable to detect the clot, then ____ principle may be used.
Chromogenic
Advantages & Disadvantages of Detection Systems of Hemostatic Analyzers
Advantages:
* No interference from specimen lipemia or bilirubinemia (icterus)
* Ability to use specimen and reagent volumes as small as 25L in some instruments
* Able to detect weak clots
Disadvantages:
* Reliance on the integrity of the entire coagulation cascade
* Inability to observe graph of clot formation
Mechanical
Advantages & Disadvantages of Detection Systems of Hemostatic Analyzers
Advantages:
* Good precision (good reproducibility of results)
* Increased test menu flexibility and specimen quality information when multiple wavelengths are used
* Ability to observe graph of clot formation with some instrumentation
Disadvantages:
1. Interference from lipemia, hemolysis, bilirubinemia and increased plasma proteins
2. May not detect short clotting times owing to long lag phase
3. May not detect small friable clots that are translucent
Photo-Optical
Advantages & Disadvantages of Detection Systems of Hemostatic Analyzers
- Resolves the limitations of both mechnical and phot-optical detection method
Advantages:
* Ability to measure proteins that do not clot
* More specific than clot-based assays
* Expanded menu options to replace clottable assays affected by pre-analytical variables such as heparin, thrombin inhibitors, or FXa inhibitors
* Most automated systems now have cost-effective chromogenic capabilities
Disadvantages:
* Limited by wavelength capabilities of some instruments
* May need large test volume to be cost-effective
Chromogenic
Advantages & Disadvantages of Detection Systems of Hemostatic Analyzers
Advantages:
* Ability to automate tests previously available only with manual, time-consuming methods, such as enzyme-linked immunosorbent assays
* Expanded test menu capabilities
Disadvantages:
* Limited number of automated tests available
* Higher cost of instruments and reagents
* May need to have additional instruments available to run routine tests in laboratories without automated coagulation analyzers that have random access capability
Immunologic
Advantages & Disadvantages of Detection Systems of Hemostatic Analyzers
Advantages:
* Ability to measure antigen-antibody reactions for proteins present in small concentrations
Disadvantages:
* Limited number of tests available
* Higher cost of reagents
* Need for special staff training
Nephelometric
- Use of electrode probes
- One of the earlierst methods that came about using the mechanical end-pointassay
- Principle: Uses KC 4/Start 4 Magnetic Sensor
Fibrometer
- Uses magnetic steel ball
- The alternating movement will cause the bead in the cuvette to do a pendular movement from left to right
- As soon as coagulation begins to be activated, the movement of the bead slows down
- The instrument will detect that and will wait for the next movement of the bead and will show a constant decline of the height of pulses
- Very reliable but there is no light that is used for the detection of the clot
KC 4/Start 4: Magentic Sensor
- Uses a magnetic sensor with a steel ball that oscillates in the arc og the cuvette
- Movement is monitored continuously within a magnetic field
- As the specimen clots, viscocity rises and the movement of the steel ball is impeded
- Variation in amplitude stops the timer
- The interval is the clotting time of the plasma
Mechanical End-point Assay
- Makes use of a light source
- Polychromatic light is focuses by a collimator and filtered to transmit a selected wavelength
- Monochromatic light is transmitted through a fiber optic mechanism and focuses the light on the cuvette where the sample is
- As fibrin clot forms, opacity increases, light scatters causing an increase in OD and the intensity of light reaching the sensor decreases
- Determines the difference of baseline OD to final OD that is recorded by the photodetector over the period of clot formation
- When OD rises to a predetermined variance from the baseline, the timer stops
- Advantage: You are able to visualize the clot waveform.
Photo-optical End-point Assay
Please study yung clot waveform in better detail, huhu di ko siya kaya icards na maayos.
So sorry </3
- A modification of the photo-optical end-point where the 90-degree or forward angle light scatter is used for measurement
- Fibrin polymers increases the light scatter
- A light emitting diode produces an incident light at approximately 600 nm
- A photodetector detects variations in light scatter at 90-degrees
- Also used for immunoassay detection of clotting factors
- Can perform both photo-optical and mechanical principle
Nephelometric End-point Assay
- State-of-the-art photometric module
- 4-simultaneous wavelength reading
- Blue dye to guarantee reagent dispensing
Optical Coagulation Detection
- The original Amelung method
- Reference method since the 70s
- No optical interferences from lipemic, iceric or hemolysed samples
Mechanical Coagulation Detection
Match the following.
- Clot Waveform Analysis
- No need for HIL sorting and result provided even with turbid sample
- The best of both technologies
A. DT 100
B. Mechanical
C. Optical
- C
- B
- A
Pathologic development of blood clots in the veins or arteries
Thrombosis
- Agents used to prevent clot formation
- Examples: Anticoagulants, antiplatelet drugs
Antithrombotics
What are the three (3) categories that antithrombotic drugs can be categorized into?
- Heparin (Anticoagulant)
- Coumadin (Anticoagulant)
- Aspirin (Antiplatelet drug)
- One of the most significant cases used to run for anticoagulation
- Also known as venous thromboembolism (VTE) - includes superficial and deep vein thrombosis (DVT) and pulmonary embolism (PE)
- Clots are formed in veins starting from the legs where the diameter of BVs are much larger
- These clots will continue to follow the circulation until they get to the blood vessels of smaller diameter and they end up in the pulmonary area which will trigger pulmonary embolism
Venous Thromboembolic Disease
What are the four (4) different treatments for VTE?
- Intravenous Standard Unfractioned Heparin (UFH)
- Subcutaneous Low-Molecular Weight Heparin (LMWH)
- Subcutaneous Synthetic Pentasaccharide (Fondaparinux), or one of the DOACs
- Vitamin K Antagonist Warfarin Sodium (Coumadin)
- Judicious use of anti-thrombotic therapy
Antithrombosis
It is very important to monitor the levels of heparin/coumadin as anticoagulants because we want to avoid what two (2) phenomena?
- Overdose
- Inadequate Dosages
This phenomena is critical and leads to emergency department visits for uncontrolled bleeding.
Overdose
This phenomena leads to secondary (repeat), often fatal, thrombotic events dosages and half-lives differ among the antithrombotics because of variations in formulation and metabolism.
Inadequate dosages
Coumadin for oral anti-vitamin K targets what?
Vitamin
UFH, LMWH, and Fondaparinux acts as inhibitors to what?
Activated Factor X
In cases of Heparin-induced Thrombocytopenia, what two drugs can be used and monitored through aPTT?
Argatroban and Bivalirudin
Match the following.
- Aspirin
- Clopidogrel, Prasugrel, Ticagrelor
- Epitifibatide, Abciximab, Tirofiban
A. VerifyNow Aspirin, AspirinWorks, Platelet Aggregation
B. VerifyNow GPI
C. VerifyNow P2Y
- A
- C
- B
Includes:
* Acute Myocardial Infarction (AMI)
* Ischemic Cerebrovascular Accident (CVA, Stroke)
* Transient Ischemic Attack (TIA)
* Peropheral Arterial Occlusion (PAO) or Peripheral Artery Disease (PAD)
Arterial Thrombosis
Arterial Thrombosis is managed by what two (2) treatments?
- Anticoagulants
- Antiplatelet Drugs
What is the preferred unit of reporting for prothrombin time when monitoring oral anti-vitamin K therapy?
A. Time in seconds
B. Percent Activity
C. INR
D. Ratio
C. INR
Used to:
* Avoid abnormal clot formation
* To stop the extension of an abnormal clot into the blood vessels
* Prevention and treatment of thrombosis
Anticoagulant Drugs/Anticoagulant Therapy
For prophylaxis and treatment of venous and arterial thrombosis
OAVKT (Oral Anticoagulant Vitamin K Therapy??? IDK)
Reduces the levels of factors II, VII, IX, and X, as well as Protein C and Protein S
Oral Anticoagulant Therapy
It is more sensitive in monitoring coumadin and warfarin because of the arrangement of the cascade.
Prothrombin Time
Please study yung Vitamin K Action on the Prothrombin Group.
Thanks pi.
This zone indicates that if the patient is taking the right dosage just to control enough the availability of Vitamin K dependent coagulation
Therapeutic Zone
T or F: Prothrombin time when performed is solely reported in INR.
F (it can be reported in seconds and percent activity as long as it is not used in monitoring patients taking coumadin and warfarin therapy)
Match the following.
- For patients under prevention and treatment of venous thrombosis and pulmonary embolisms.
- For patients with mechanical prosthetic valves or recurrent thrombosis and embolisms with OAT.
A. Therapeutic Range: 2 - 3 INR
B. Therapeutic Range: 3 - 4.5 INR
- Was used prior to the introduction of INR unit
- What was did in the laboratory to establish percent calibration
PT Results in % Activity
- An important part of INR calculation
- INR = (PT/MNPT)^ISI
- Not the control value run per day
- Determined by running fresh plasma samples from at least 20 healthy individuals
- Best tested over several days to take account of day-to-day variation of measurement
Mean Normal PT
Findings and Recommendations for Therapeutic Monitoring
T or F: PT monitoring of warfarin treatment is very precise when expressed as ratio (Patient PT/Control PT).
F (imprecise)
Findings and Recommendations for Therapeutic Monitoring
- These differences in ____ contributed to clinically important differences in oral anticoagulant dosing and were responsible for excessive and erratic anticoagulant.
Thromboplastin Responsiveness
The way to convert time in seconds to % activity is to use a?
Calibration Curve
Findings and Recommendations for Therapeutic Monitoring
- These shortcomings in the PT monitoring stimulated the development of the ____ for monitorign Oral Anticoagulant Therapy.
INR Standards
Findings and Recommendations for Therapeutic Monitoring
- The adoption of this standard improved the safety and ease of monitoring ____
OAVKT
Findings and Recommendations for Therapeutic Monitoring
- According to CAP Survey, implementation of the INR standards in the US rose from ____ to ____ between 1991 and 1997
21% to 97%
What four (4) factors affect the results in long-term warfarin therapy?
- Changes in diet
- Concurrent medication changes
- Poor patient compliance
- Alcohol consumption
- May be used as an alternative to the PT/INR System
- Therapeutic range is typically about 20% to 40% of normal factor X activity
- Useful when the PT is comprised by a lupus anticoagulant, a factor inhibitor, or a coagulation factor deficiency
Chromogenic Factor X Assay
- In case of heparin overdose
- When CPB (Cardio-Pulmonary Bypass) is discontinued, UFH (Unfractioned Heparin) anticoagulation must be quickly reversed
- Protamine Sulfate is administered by IV
- May cause a delayed form of HIT (Heparin-Induced Thrombocytopenia)
- Platelet counts are also monitored due to HIT
Reversing Heparin
Drugs given to control the activity of the platelets
Antiplatelet Drugs
- Used by cardiologists to infuse GPIs to maintain vascular patency during PCI for intracoronary stent placement and angioplasty
- Intravenous glycoprotein IIb/IIa inhibitors
- PT, PTT, ACT, Hemoglobin, Hematocrit, and Platelet Count are determined to detect any hemostatic or hematologic abnormality
- GPI efficacy may be measured using the Multiplate Analyzer or VerifyNow IIb/IIa Assay
Intravenous Antiplatelet Drugs for Percutaneous Coronary Intervention (PCI)
Oral Antiplatelet Drug to Reduce the Incedence of Arterial Thrombosis
- Prescribed at 81 to 325mb/day
- Prevents: Secondary thrombotic events in patients with stable/unstable agina, AMI, transient cerebral ischemia, peripheral vascular disease, or stroke
Aspirin
Oral Antiplatelet Drug to Reduce the Incedence of Arterial Thrombosis
- Prescribed at 75mb/day
- Is a prodrug and patients have varying dose responses
- CYP2C19 liver enzyme
Clopidogrel
Oral Antiplatelet Drug to Reduce the Incedence of Arterial Thrombosis
- A prodrug whose main active metabolite is formed via CYP3A4 liver enzyme
- Provided in 90 mg tablets
- Susceptible to drug interactions
Ticagrelor
Oral Antiplatelet Drug to Reduce the Incedence of Arterial Thrombosis
- A prodrug that is converted in the lvier via several cytochrome P450 pathways
- Half-life is about 7 hours
- Affected by drug interactions
- Carries a higher risk of bleeding
- Associated with an increased risk of solid tumors
Prasgurel
VerifyNow
- Uses light transmittance aggregometry to test for responses to aspirin, clopidogrel, prasugrel, ticagrelor, and the GPIs
- Uses arachidonic acid as its antagonist
VerifyNow System
Match the following.
- Uses ADP
- Uses thrombin receptro-activating peptide (TRAP)
A. VerifyNow IIb/IIIa
B. VerifyNow P2Y12
- B
- A
- Used to assess aggregation
- From HELENA
- 1 unit consists of 4 measuring chambers in order to see if the platelets are responding to the antagonist, the curve will rise
AggRAM Analyzer
- Platelet aggregometer
- Fully automated platelet analyzer
- Contribution of Bankanalyser
- Yo ucan see thinsg visually responding and also get quantitative results
Thrombomate XRA
- Detects clot formation for whole blood clotting
- Not high volume assay
- Provides much more information about the whole blood clotting process
Viscoelastic Hemostasis Test Measurement (Whole Blood Clotting Assay)
Please read/watch the lecture huhu, ang hirap neto.
Thanks pu.
