INTRODUCTION:
The use of synthetic calibrator matrices in Immunoassays have been previously
reported.1
Manufacturers of Immunodiagnostic kits instruments reagents, etc. have regulatory
requirements that involves providing controls material to the end user that will assure
consistently precise and accurate potential results. Calibrators also sometimes refer as
standards and controls with long term, preferably stable at ambient temperature and
appropriate but accurate concentration of analyte and accurate assignment and target
values are absolutely critical for this purpose. Unfortunately, even though calibrators
prepared from human sources, lot to lot variation in their makeup make it very difficult
if not impossible to make them behave as patient samples. These variations or properties
are sometimes referred to as matrix effects.
Since non-isotopic immunoassay introduced in the early 70s, a number of manufacturers
have made several attempts to manufacture or synthesize various matrices, that includes
human blood, animal serum and other endogenous or non-endogenous proteins, or protein-like
substances in various configurations to mimic human serum. Although some manufacturers
were able to synthesize formulation that works well for one or two analytes, lthey were
unable to make universal "cocktail" which could be substituted as "matrix
solution." In addition, they encountered another problem associated with protein
based calibrators as shown in Table #1'. The presenter and author of this article has
developed synthetic matrices under SeraSub~ and UriSub~ which are substitute matrices for
serum and urine respectively.'
MATERIALS AND METHODS:
The test kits used were obtained directly from the
manufacturers, that included Abbott Laboratories, Chicago, IL,
Technicon Corp., Tarrytown, NY, Becton Dickinson, Orangeburg, NY.
The salts and other buffer material was obtained from Sigma
Chemicals, St. Louis, MO.
DISCUSSION:
SeraSub(r) is nonbiological organic polymer material which can be used as a base for
either standards (calibrators) or controls. Both SeraSub(r) and UriSub(r) are stabile at
ambient temperature (20-330C) for at least five years if not more. More interestingly,
when analyte such as hapten or antigen is added to the SeraSub(r) or UriSub(r), the
stability at room temperature increases tremendously.
We undertook extensive study to determine the usefulness of SerSub(r) and UriSub(r) and
their stability after the particular analytes were spiked in. The Study Protocol was as
follows: The standard calibrates were prepared using SerSub(r) or UriSub(r) as required.
Each container was exposed to 450C, 370C ambient temperature (20-300C) +40C and -200C for
one week (to simulate shipping condition) and then stored at ambient temperature for the
entire study period which lasted from 11 months to 36 months. The analysis was performed
periodically. In all cases it showed no degradation of the product during the study
period. The data is shown in Tables 2 to 5.
The data presented in Table No. 2 and Table No. 3 include comparison of patient sample
results calculated based on standards generated using SerSub(r) and standards supplied by
the manufacturers. The study was conducted after at least four weeks of ambient storage of
the calibrators made using SeraSub(r). All the other components were stored according to
the manufacturer's guidelines.
SUMMARY AND CONCLUSION:
Synthetic matrices reported in this paper not only offer stability, compatibility,
safety but several other features which are listed below:
1. Safety: Because the material is nonbiological, it does not have the HIV, hepatitis,
and other infective possibilities of HSA and other materials. Benefits for the
manufacturer are safety, reduced testing and handling cost, and reduced liability.
Benefits for the laboratorian are safety and reduced handling cost.
2. Multianalyte calibrators and controls: Because the material has no endogenous
substances, it is possible to produce multianalyte calibrators and controls and simple
"labeled subsets" as needed without concern about interferences. Therefore, one
multianalyte lot will suffice for a complete range of "subset" standards (i.e.,
B12/folate, thyroid panel, or complete panel as required), and the same matrix can be used
for either standard or control without stripping.
Benefits for the manufacturer are:
Single stock, various labels
Interchangeable material for standards and controls
Ease of manufacture of full line
Lower cost of manufacture and quality control
Benefits for the laboratorian are:
Both subset and complete standards or controls can be the same
Reduced cost
Reduced variability
3. No endogenous anal ytes: Because the material is nonbiological, it has no endogenous
interfering substances. Hence, the matrix has the following attributes:
No "stripping of unwanted substances" required Easy to make "0"
standard
Very low (NSB) nonspecific binding
Benefits for the manufacturer are reduced quality control efforts, improved yield, and
reduced cost. The benefit to the laboratorian is simplified user calculations or
adjustments.
4. Stability without lyophilization: Because it is nonbiological, the material is not
susceptible to degradation due to elevated temperature exposure during typical handling.
This produces the following benefits. For the manufacturer:
No lyophilization process required
No special packaging
Stable in liquid form
Storable at room temperature
Shippable at ambient temperature
Large single lots possible
Reduced cost of manufacture storage,
packaging, quality control, and shipping
For the laboratorian:
Ready to use without reconstitution, reducing errors and improving ease of use
Easy to store
Reduces lot change problems
5. Consistent properties: Because the material is not derived from a living host, it
can be produced in large quantities with consistent properties, therefore eliminating
lot-to-lot variations commonly associated with biological matrices. Benefits to the
manufacturer include higher yield, lower cost to produce, less inspection required, and
the fact that it takes only 1/2 hour to produce a homogeneous solution. The principal
benefit to the laboratorian is continuity of quality control due to consistency between
lots.
TABLE 1
Disadvantacres of Protein-Based Calibrators
1. Potential shortages in the supply of human blood products due to fewer donors, high
donor rejection rates, and improved whole blood utilization resulting in fewer outdated
blood units becoming available for in vitro use.
2. Large and expensive inventories must be maintained to minimize screening and
validation expenses.
3. The relatively high cost of human and animal products, particularly of purified
blood fractions.
4. The variable and frequently diminishing endogenous enzyme levels and their
unpredictable effect on calibrator performance.
5. Protein matrices are excellent growth media requiring aseptic manipulation and/or
use of effective antimicrobial agents.
6. Presence of viruses and infectious agents in some approved lots due to the
limitations of current testing procedures 2.
7. Potential contamination with, and the need for testing for, pathogenic or
nonpathogenic microorganisms and viruses, including hepatitis virus and HIV.
8. Potential contamination of protein matrices with unacceptably high levels of
endogenous analytes, necessitating in-house processing or purchase of more costly
"stripped" protein matrices.
9. Inherent instability of proteins or protein solutions requiring storage at freezer
temperatures.
10. Lot-to-lot variability.
