R.

Gagnon

August 2009

Contents
1. Introduction- What is a clot signature curve.. 3
2. Parts of the clot signature curve.. 3
3. What to examine in the curve segments 4
4. What can affect the curve appearance?................................. 5
5. Curve shape characteristics and possible causes.... 6
6. Comparison of good and bad curves... 7
7. How to display a curve on the ACL ELITE / ELITE PRO..9
8. Examples of clot curves .. 11
9. Data Reduction flow chart for PT/APTT/TT ..19
9. Analytical Error and Warning Codes... 20

The Clot Signature Curve; many of you have heard of it but what do you really know about it?
This document is intended to provide answers to the following questions about clot signature
curves:
-

What is a Clot Signature Curve?

How is a Clot Signature Curve created?
What things will impact the formation of the Clot Signature Curve?
How can I use the Clot Signature Curve in my laboratory?

What is a Clot Signature Curve?

A clot signature curve is a visual representation of the coagulometric or absorbance reaction
taking place in a cuvette on the analyzer. A clot signature curve can easily be displayed for all
results on the ACL ELITE / ELITE PRO analyzers.
The clot signature curve can be a valuable tool for investigating abnormal or questionable results.
In addition, it can also be used to investigate warnings or errors during sample analysis.

Y- Axis

X- Axis

Parts of the Clot Signature Curve

-

Y Axis: Representation of nephelometric (light scatter) reading. As fibrin forms it

increases the amount of light scattered. On the ELITE this is represented as a ratio
between the sample and reference (Wash-R) times 100.
X Axis: Increase in Time going from left to right
Delay: Sample monitoring begins at time zero. During the Delay time the sample
and reagent are initially mixed. Data acquired during this period is not used in the
data reduction calculations.

Baseline: Commences the start of the data acquisition. This segment begins after
the delay time; after the sample and reagent are mixed. During this segment little
change in the optical reading will occur until the fibrin clot begins to form. The
baseline is generally short for normal clotting reactions, and will be longer for
abnormal samples.
Acceleration: This segment represents the fibrin formation period. During this
segment the optical change will be rapid resulting in the steep slope rise. Slow clot
formation will be displayed as a curve with a shallow slope.
Deceleration: Immediately follows the Acceleration segment and represents the
decrease in the rate of clot formation. During this segment the available fibrinogen
will have been converted to fibrin and thus the optical change and reaction slope
plateaus off. Weak clots from samples with low fibrinogen may exhibit a slight
decrease during this segment as the clot formed is fragile.
Endpoint: The point at which the data acquisition stops.
Delta: Change in absorbance between the baseline and endpoint phase

What to examine in the curve segments

The clot curve above has the characteristic Sigmoidal, or slanted S shape. The S shape
results from the baseline, acceleration and deceleration segments.
The areas to investigate within each of these segments include the following:
1. Baseline
- Starting point reading on the Y axis
- End point reading on the Y and X axis
- Stability: Is the baseline segment smooth or bumpy?
- Slope: Is the baseline rising rapidly?
2. Acceleration segment
- Starting point reading on the Y and X axis
- End point reading on the Y and X axis
- Slope: Is it a steep rise or low, gradual rise?
- Is the Acceleration segment line smooth or bumpy?
3. Deceleration and Plateau segment
- Starting point reading on the Y and X axis
- End point reading on the Y axis
- Stability: Is this segment smooth or bumpy?
- Slope: Is the plateau flat, rising or falling?

What can affect the Curve appearance?

The clot/reaction curve results from the interaction of the sample and reagent in conjunction with
the analyzer (temperature, mixing, incubation time, etc). Anything that can affect these items
could impact the clot curve.
-

Sample quality
o Improper collection
o Clotted sample
o Improper sample storage
o Inadequate centrifugation
o Over anti-coagulated (elevated hematocrit)
o Under anti-coagulated (low hematocrit)
o Frozen sample thawed incorrectly
o Sample Age

Reagents
o Improper reconstitution
o Expired reagents (onboard stability and / or lot expiration)
o Incorrect placement on analyzer
o Lack of stir bar usage when required
o Reagent contamination

Instrument Issues
o Sample and Reagent Probe condition
o Sample and Reagent Aspiration component condition
o Spills / Dust in the reading area
o Environmental surroundings electrical interference
o Contamination
o Dirty cuvettes
o Damaged optical components

Clinical Condition of the Patient

o Factor level deficiency
o Disseminated Intravascular Coagulation (DIC)
o Sepsis
o Excessive lipemia
o Liver Disease
o Anticoagulant therapy
o Drug interference

Curve Characteristics (shape) and possible causes

Curve Characteristics
- Flat Curve (Indicative of no clot formation)

Possible Cause
- Lipemia or other interfering substance
masking clotting activity
- Low Fibrinogen
- Excessive Anticoagulant
- Reagent Quality
- Sample Quality
- Instrument fluidic handling issue

Shallow Acceleration Segment

- Slow clot formation due to slow conversion of

fibrinogen to fibrin (similar to a flat curve)

Small changes in Raw Counts from

Baseline to Endpoint

- If extremely small Optical change and very short

clot time, noisy baseline causing false result
- Low Fibrinogen concentration

Large Change in Raw Counts from

Baseline to endpoint.

- High Fibrinogen concentration

Endpoint drops towards Baseline

- Unstable clot

Endpoint rises

- Clot formation is still progressing

Acceleration segment seen but no Endpoint - Clot formation did not occur within the
Acquisition time

Long Baseline

- Prolonged/impaired clotting mechanism

Comparison of good and bad curves

800
700
600
500
400
Note the Bumpy Baseline Starting around 300
on the Y Axis

300
200
100

Note Smooth Baseline Starting around 220 on the Y Axis

10

15

20

25

30

35

40

45

50

800
700

This curve has an acceleration

segment with a good slope

600
500
400
300
200

This curve has an acceleration segment with a

shallow slope and increasing plateau segment.

100
10

15

20

25

30

35

40

45

50

This curve has a nice stable plateau

800
700
600
500
400
300

This curve has a decreasing baseline and a

plateau segment that decreases over time
and ends up lower than the starting point.

200
100
10

15

20

25

30

35

40

45

50

How to Display a Curve on the ACL ELITE / ELITE PRO:

-

Select the desired sample from the main results (database) screen and press the Detail
icon

Select the desired result for which you wish to view the clot curve and press the Detail
icon

The clot curve will then be displayed

Example of a printed Clot Curve from the ELITE / ELITE PRO Analyzers

Important Summary Notes:

When you look at a clot curve always remember to observe the following:
1. The scale of the Y axis on the graph, see if it is a narrow or wide optical reading
band.
The system always maximizes the curve to fit the entire window. Observation of the
Y axis readings is the only way to determine the delta between the baseline and
endpoint.
2. The shape of the curve, it should generally have the 3 segments: baseline,
acceleration, and plateau.
3. What are the initial and endpoint readings?
4. Note the vertical line which denotes the clot point.

10

Example Clot Curves for PT and APTT

This section of the document will present PT and APTT clot curves from the ELITE / ELITE PRO
systems. The examples will include discussion information on the shape of the curve along with
any flagging that occurred

1. Normal PT Curve

This PT curve illustrates what a normal sample for PT should look like. This curve has a classic
baseline, acceleration and plateau phase. Also note the Y axis scale illustrates a good delta
between the baseline and endpoint. The Clot time was 12 seconds for this sample.

11

2. Extended PT Curve

This sample illustrates a typical clot curve for a sample with an extended PT clotting time. On this
sample you will note:
- Long baseline phase
- Standard acceleration phase
- Standard deceleration/plateau phase
- Acceptable delta between baseline and endpoint based upon the Y axis readings
The reading for this sample is taken at 75.2 seconds.

12

3. PT Delta Error

This sample illustrates a PT clot curve with an unstable decreasing plateau phase
- The reaction starts out as a normal S shaped curve, however after the maximum
reading is achieved the reaction begins to decrease.
- Note the Y axis delta ranges from 43 to 89 which is good
- This is indicative of a clot that is not stable.
This sample resulted in a Failed reading with an error 11 (curve dropping after reaching its
maximum) This sample should be repeated.

13

4. PT Error due to Reagent Issue

This sample illustrates what appears to be a standard clot curve. When viewing the curve note:
-

Y axis scale ranges from 14 to 26.6 resulting in a low delta for the curve
Y axis reading is starting off very low compared to a normal PT
Low reactivity resulting from reagent issue (contamination, degradation)

This sample resulted in a Failed reading, error 12 (coag error), for the PT. Sample should be
repeated. If failure occurs a second time then controls should be run. If the controls are not
within the specifications, new reagent should be prepared. If the controls are acceptable, then the
sample should be investigated for possible low fibrinogen concentration.

14

5. Normal APTT Sample

This sample illustrates a typical clot curve for a sample with a normal APTT clotting time. On this
sample you will note:
- Standard baseline phase for APTT
- Standard acceleration phase
- Standard deceleration/plateau phase
- Acceptable delta between baseline and endpoint based upon the Y axis readings
The reading for this sample was 29.8 seconds.

15

6. Extended APTT Sample

This sample illustrates a clot curve for a sample with an extended APPT clotting time. On this
sample you will note:
- Extended baseline phase for APTT
- Standard acceleration phase
- Commencement of the deceleration/plateau phase
- Acceptable delta between baseline and endpoint.
This sample resulted in a reading of 118.2 seconds for the APTT.

16

7. APTT Error

This sample illustrates a clot curve for an APTT sample run in the standard mode. On this
sample you will note:
- Lack of the standard S shape to the reaction curve
- Readings on the Y axis range from 45 to 50 reflecting a lack or reactivity and therefore
a low delta
- A rising baseline appears to be present followed by what appears to be the start of the
acceleration phase.
- This sample should be rerun using the extended mode
The reading for this sample resulted in a Failed result from error 31 (minimum delta not met).
This clot curve was obtained from a patient on heparin

17

8. Lipemic Sample APTT

This sample illustrates a APTT clot curve for an extremely lipemic sample
- Note the Y Axis reading. The reading starts around 340. For a normal sample this is
generally in the range around 40.
- The curve does not present the classic S form.
This sample resulted in a Failed reading with an error 31 (minimum delta not met)

18

Data Reduction for PT and APTT

Note:
The first derivative is defined as the maximum velocity of the curve.
The second derivative is defined as the maximum change in velocity (acceleration) of the curve
or the fastest rate of fibrin formation.

19

Analytical Error and Warning Codes

The tables below include codes and flags that may be associated with curves/ results
Error code - 5
Meaning

Optical failure

Cause

ADC saturation (signal above 9.5 V at the end of the clotting

curve).

Flags

Results

Error 5 instead of the result.

Remedial
Action

Possible high Fibrinogen concentration. Dilute the sample

1:1 with factor Diluent and repeat the test.

Error code 6
Meaning

Not coag

Cause

First threshold not passed.

Flags

Results

Error 6 instead of the result.

Remedial
Action

Sample does not clot within the acquisition time.

Repeat the test in extended acquisition time.

Error code 7
Meaning

Coag error

Cause

Second threshold not passed.

Flags

Results

Error 7 instead of the result.

Remedial
Action

Sample clot curve is noisy and does not give a normal clot
signal within the acquisition time. Repeat the test in extended
acquisition time.

Error code - 8
Meaning

Coag error

Cause

Delta time between the two thresholds is higher than the

selected value.

Flags

Results

Error 8 instead of the result.

Remedial
Action

Possible non-phasic clotting curve. Review the clot curve.

Possible sample interference with the clotting reaction.

20

Error code - 9
Meaning

Coag error

Cause

Initial slope of the reaction curve is higher than the

selected value.

Flags

Results

Error 9 instead of the result.

Remedial
Action

Possible bi-phasic clotting curve. Review the clot

curve. Possible sample interference with the clotting
reaction.

Error code - 10
Meaning

Coag error

Cause

Final slope of the reaction curve is higher than the

selected value.

Flags

Results

Error 10 instead of the result.

Remedial
Action

Unstable endpoint of the clotting curve. Review the clot

curve. Possible sample interference with the clotting
reaction. Repeat the test in extended acquisition time.

Error code - 11
Meaning

Final delta error is a check that the curve is not

dropping too much after reaching its maximum reading

Cause

Final delta of the reaction curve (maximum abs reading

final abs. reading) is higher than the selected value.

Flags

Results

Error 11 instead of the result.

Remedial
Action

If this is a nephelometric reaction, it may be an

indication of an unstable endpoint in the clotting curve.
Review the clot curve. Possible sample interference
with the clotting reaction. Repeat the test in extended
acquisition time.
If this is an absorbance test, it may be an indication of
an absorbance value outside the specified limit.

21

Error code - 12
Meaning

Coag error

Cause

Maximum peak of the first derivative is below the

selected limit value.

Flags

Results

Error 12 instead of the result.

Remedial
Action

First derivative peak is not significant enough to

indicate a real clotting reaction point. Review the clot
curve. Repeat the test in extended acquisition time.

Error code 13
Meaning

Coag error

Cause

Maximum peak of the second derivative is below the

selected limit value.

Flags

Results

Error 13 instead of the result.

Remedial
Action

Second derivative peak is not significant enough to

indicate a real clotting reaction point. Review the clot
curve. Repeat the test in extended acquisition time.

Error code - 14
Meaning

Offset error (delta algorithm)

Cause

Offset value is greater than First part value defined in

calculation setup section of test definition or below
scale range low limit.

Flags

Results

Error 14 instead of the result.

Remedial
Action

Review the clot reaction curve Y axis scale to

determine if value is low or high (turbid). Rerun
Sample.

Error code - 30
Meaning

Offset error (delta algorithm)

Cause

Offset of the initial part of the curve is below the

selected limit value.

Flags

Results

Error 30 instead of the result.

Remedial
Action

Initial reaction turbidity is relatively low. Review the clot

reaction curve. Check integrity of reagents, and make
sure no bubbles are present

22

Error code - 31
Meaning

Minimum Curve Delta not met

Cause

The total delta of the reaction curve is less than the

limit specified in the test setup. (Reaction curve is flat,
and clot formation may not have occurred)

Flags

Results

Error 31 instead of the result.

Remedial
Action

Review the curve and rerun the sample. Sample may

have an extended clotting time.

Noisy Baseline/Reaction Curve (Error 32/33)

Meaning

Reaction curve baseline readings are erratic

Cause

Interference in reaction readings

Flags

Q - Noisy Baseline/Reaction Curve displayed in

Warning List box when clot curve is displayed

Results

Error 32/33 instead of the result

Remedial
Action

Review Curve and repeat sample

23