Intended Use:
To quantitate total human
Apolipoprotein A1 (Apo
A1)
Principle of
Procedure:
Solid phase
capture sandwich ELISA assay using a
microwell format.
Shelf Life:
The expiration date for the package and each component is stated on the
label(s).
Store all components at 2°- 8° degrees C except for the standard, which should be stored at -20oC.
Patient and Standard Dilutions:
*Dilute the 15X wash buffer provided 1:15 using one part wash buffer concentrate and 14 parts reagent grade water.
Dilute each serum, plasma
or tissue culture fluid specimen to
be tested 1:10,000
with diluted wash buffer to form a final 1:10,000 dilution.
Prepare serial two fold dilutions of the human Apo A1 standard: Neat (N), 1:2,
1:4, 1:8 etc. using diluted wash buffer. Use diluted wash buffer alone as
the blank or zero control well.
Materials Supplied:
Anti-Human Apo A1 coated microwell strips 12 x 8 with
plastic frame
HRP conjugated affinity purified goat anti-Apo
A1 - 12mL
**Apo A1 standard (pre-diluted 1:10,000) - 1 ml
TMB/peroxide substrate color developer - 12mL
Sulfuric acid termination reagent (0.5N) - 12mL
15 X
Wash buffer concentrate - 2x60mL
Limitations of the Procedure:
No single assay should be used as the only basis for
arriving at a diagnostic conclusion.
For research use only.
Dynamic Range:
9.375 mg/dl-600mg/mL
Reproducibility:
C.V. 6% - 10% depending upon the region of the standard curve.
Assay Procedure:
*Allow each reagent to reach room temperature before use.
1. Add
100uL of diluted specimen or standard to each microwell.
2. Incubate
at room temperature for 2 hours
3. Decant
and wash each microwell four times with wash buffer (dilute buffer 1:15 with
reagent grade water)
4. Add
100uL of HRP conjugated goat anti-Apo A1 to each well
5. Incubate
at room temperature for 2 hours
6. Decant
and wash as in "step 3".
7. Add
100uL of TMB/peroxide substrate and incubate at room temperature for 30 minutes
8. Terminate
the reaction with 100uL of 0.18N sulfuric acid
9. Zero
the microwell reader at 450nm using the specimen diluent zero control well
10. Determine
the optical density (O.D.) of the remaining wells.
11. Construct
a standard curve using the O.D. values obtained for each of the standards
12. Interpolate
the unknowns from the standard curve
**Note: This Apo A1 Standard has been calibrated against the International Federation of Clinical Chemistry (IFCC) Standard, Lot Number 293 and has been demonstrated to recover 100% of this standard.
Typical Standard Curve:
This assay has been demonstrated to recover 100% of the Internation Federation of Clinical Clinical Chemistry (IFCC) Standard BRC-393