Adequate Internal Controls Research Articles

Extraction buffer contaminated bacterially as a cause of invalid HIV-1 viral load results on the NucliSens EasyQ ® system

Shortly after starting to use the NucliSens EasyQ ® HIV-1 V1.1 system for HIV-1 RNA load testing, the number of invalid tests per assay run gradually increased. Within five days, approximately 50% of tests showed a total lack of amplification of the calibrator and in most cases also of the HIV-1 template. According to the manufacturer's specifications, the lysis buffer and three extraction buffers remain on the automated NucliSens easyMAG™ extraction system between assay runs. Therefore possible microbial contamination of these buffers was investigated, after they had been on the automated system for approximately one week. The NucliSens easyMAG™ extraction buffer 2 yielded bacterial growth identified as Acinetobacter baumannii. After regular decontamination of the machine's tubing system with 70% alcohol and storage of the buffers at 4 °C between assay runs were commenced, invalid results due to failed internal calibrator signal occurred no longer. It is likely that bacterial contamination of the buffer was the cause of assay failure, probably due to ribonuclease (RNase) activity. Bacterial contamination of PCR systems should be added to the list of potential hazards in diagnostic virology. This experience underlines the necessity of state-of-the-art assay design incorporating adequate internal controls and calibrators.

Fighting costly health care fraud

AbstractThere is a rising tide of health care fraud in this country. And it is costing all of us upwards of $100 billion each year, adding to already skyrocketing medical costs. Many companies do not have adequate internal controls over their health care system. And even when they do, monitoring or auditing is lax. The authors explore the scope of these problems and offer practical advice on stemming the tide. © 2005 Wiley Periodicals, Inc.

Quantitation of low abundance mRNAs in glial cells using different polymerase chain reaction (PCR)-based methods

The conventional methods for mRNA quantitation such as Northern blotting or ribonuclease protection assay sometimes lack enough sensitivity to study low abundance mRNAs or to work with limited amounts of biological samples. The sensitivity of the polymerase chain reaction (PCR) linked to reverse transcription (RT-PCR) has proven useful in amplifying specific mRNAs, especially those present in low copy number. Though, the quantitation of nucleic acids by means of PCR has proven problematic. The main constraint in obtaining quantitative data is inherent in the amplification reaction. Because amplification is an exponential process, small variations in the efficiency of amplification may significantly affect the final yield of the PCR product. The variables that influence the rate of the PCR include the abundance of the mRNA present in the starting material, the concentrations of the Taq DNA polymerase, dNTPs and magnesium ions, the annealing and elongation conditions, the ramping temperatures and the formation of primer secondary structures. Moreover, with the progression of the PCR cycles, reagents are consumed and inhibitors generated, leading to non-linear synthesis of DNA. Finally, tube-to-tube variations sometimes preclude accurate quantitation. Most of the above-mentioned problems can be overcome by the choice of adequate internal controls. The present report reviews two recently developed methods for RNA quantitation, the semi-quantitative PCR and the quantitative PCR illustrated for the measurement of monoamine oxidase (MAO) A and B mRNAs and the estrogen receptor (ER) mRNA respectively, with a particular emphasis on the design of appropriate internal controls to compensate for the intra- and inter-assay variability inherent to RT-PCR.

Utility of an internal control for the polymerase chain reaction. Application to detection of Mycoplasma pneumoniae in clinical specimens.

The polymerase chain reaction (PCR) was used to amplify a 209 base-pair fragment of Mycoplasma pneumoniae DNA. The amplicon was transferred into a plasmid and a 680 base-pair piece of foreign DNA was inserted between the two amplimer sites. Plasmid DNA was added to the reaction mixture as an internal control for the polymerase chain reaction. Since the original hybridization target sites were included in this construction, one pair of amplimers could be used to amplify both the target DNA and the internal control DNA. Separation of internal control from target DNA after amplification was easily obtained on agarose gel electrophoresis. For the analysis of clinical samples with the polymerase chain reaction, the addition of internal control DNA allowed monitoring of the overall effectiveness of the amplification in each tube. With this technique approximately one-third of the tests were shown to be unsatisfactory due to technical errors or contaminating inhibitors. Adequate internal controls are necessary to avoid false-negative results with the polymerase chain reaction.

Academia Careful On Direct R&D Costs

Unlike many indirect expenses charged to federal funding agencies and garnering sensational headlines during the past year or so, universities seem to proceed very carefully, with a few exceptions, about how they spend their direct research dollars. The General Accounting Office delivered that assessment late last month during hearings of the Senate Committee on Governmental Affairs, chaired by Sen. John Glenn (D.Ohio). Because 70% of the $9 billion the government budgets annually for support of R&D at academic institutions goes to fund direct research costs, Glenn had asked GAO to ascertain whether adequate procedures, internal controls, and resources are in place and working effectively to protect the government's interests. On hearing GAO's conclusion, Glenn noted: Thankfully their findings did not show outrageous charges for most direct costs, unlike some of the recent indirect costs that have been noted in the press. Glenn had asked GAO to limit its investigation to recipients of National Science F...

Internal Accounting Controls in the Office of the Future

The law, as well as good business practice, requires adequate internal accounting controls. Modeling and machine analysis of office information systems can ensure compliance.

The Adequacy of Internal Corporate Controls

In order for a corporation's activities and con duct to be what its top management wants, there must be ade quate internal corporate controls to assure that management knows what is, in fact, going on and that management's in structions are being carried out. The antitrust electrical cases, the heavy losses suffered by General Dynamics through its Con- vair division, and Chrysler's troubles over conflict of interests among its executives are illustrations which point out the fail ure of internal systems of control. In each case, top manage ment did not have the benefit of adequate internal controls so that it could prevent illegal or improper or unfortunate events from occurring, to the detriment of the corporation. These in stances suggest the timeliness of an inquiry into the adequacy of internal corporate controls in the large American corpora tions in order to ascertain what really is "the corporation."