COMMON UNITS OF MEASUREMENTS USED IN THE CLINICAL MOLECULAR LABORATORY

The clinical molecular laboratory has become an integral part of the diagnostic testing menu. Actual viral loads for the human immunodeficiency virus (HIV), or hepatitis B or C, has dramatically changed treatment options for patients. It no longer is good enough to be able to measure the presence of the HIV antibody in patients; now treatment protocols demand to know the exact amount of virus present in the body.

The work that is performed in a clinical molecular laboratory differs from that performed in a research molecular laboratory or in a biotechnology production laboratory. It is not the purpose of this chapter to go into detail on the many calculations that are associated with research protocols or biotechnology protocols that range from calculating bacterial growth to developing unique probes, to name just a few. There are many reference books on the subject, many reliable molecular biotechnology sources, blogs, and discussion forums that can be found on the Internet, and molecular and biotechnology professional organizations that provide a forum for discussion among members. A few of the professional societies are the American Society for Biochemistry and Molecular Biology, the American Society for Microbiology, the American Society for Cell Biology, and the European Laboratory for Molecular Biology. The University of Illinois’s library has a comprehensive listing of molecular and cell biology weblinks. There is also an extensive molecular biology forum originally linked to the National Oceanic and Atmospheric Administration and the Northwest Fisheries Science Center (http://molecularbiology.forums.biotechniques.com/forums/index.php).

A list of molecular biology websites can be found at the end of the chapter.

The metric system was discussed in Chapter 3. However, in the molecular laboratory much smaller quantities of materials and dilutions are performed versus what is performed in the core laboratory. For example, if a 1 to 10 dilution was to be performed in a clinical chemistry laboratory, the dilution might be performed with 10 μL sample to 90 μL diluent. Compare that to the dilutions performed in the clinical molecular laboratory where a 1 to 10 dilution might be performed with 1 μL sample in 9 μL diluent.

Table 12–1 presents common metric units that are used in a clinical molecular laboratory.

Dilutions/Solutions Used in the Clinical Molecular Laboratory

Buffers are used in the clinical molecular laboratory for a variety of tests. For example, if samples are tested by electrophoresis, then buffers are integral in the electrophoresis technique. Common buffers used are Tris borate buffer, TE buffer, STET buffer, and maleic acid buffer. The calculations used are generally C1V1 = C2V2 calculations to form a working buffer from a stock solution. What makes the difference between the molecular laboratory and the main laboratory is the volumes used. The clinical molecular laboratory in general uses much smaller quantities of patient sample and reagents than what is commonly used in the main clinical laboratory. Ethanol is also commonly used in many molecular techniques, many times at a 70% ^v/v concentration.

Example 12–1

A medical laboratory scientist student needed to prepare a 70% ^v/v ETOH solution to be used in a hepatitis C molecular assay. The stock ETOH was at 100% ^v/v. and 100 mL of the 70% ETOH solution was needed. How did the student prepare this solution?

The C1V1 = C2V2 formula is used to solve this problem.

C1 = stock concentration (100%)

V1 = volume of stock needed

C2 = 70%

V2 = 100 mL

Using C1V1 = C2V2 the following equation is derived:

(100)(X)=(70)(100)100X=7000

Solving for X yields:

Therefore, 70 mL of the stock ETOH is added to a 100 mL volumetric flask, and 30 mL deionized water is added to form the 70% ^v/v ETOH solution.

CONVERSIONS COMMONLY PERFORMED IN A CLINICAL MOLECULAR LABORATORY

When double-stranded DNA (dsDNA), single-stranded DNA (ssDNA), or oligonucleotides are extracted, their quantity may be expressed in molar terms. Table 12–2 contains the molecular weight of DNA, RNA, and oligonucleotides in terms of picograms per picomole.

To convert dsDNA that is in terms of micrograms per milliliter to picomoles per microliters, use the following formula:

n = number of nucleotides

X = number of micrograms per milliliter of DNA that you have

To convert from picomoles per microliter DNA to micrograms per milliliter DNA, use the following formula:

XpmoldsDNAμL×660pgpmol×1000μL1mL×1μg106pg×n=μg/mLdsDNA

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