Analytical validation of HER2DX genomic test for early-stage HER2-positive breast cancer.

BACKGROUND: HER2DX, a multianalyte genomic test, has been clinically validated to predict breast cancer recurrence risk (relapse risk score), the probability of achieving pathological complete response post-neoadjuvant therapy (pCR likelihood score), and individual ERBB2 messenger RNA (mRNA) expression levels in patients with early-stage human epidermal growth factor receptor 2 (HER2)-positive breast cancer. This study delves into the comprehensive analysis of HER2DX's analytical performance. MATERIALS AND METHODS: Precision and reproducibility of HER2DX risk, pCR, and ERBB2 mRNA scores were assessed within and between laboratories using formalin-fixed paraffin-embedded (FFPE) tumor tissues and purified RNA. Robustness was appraised by analyzing the impact of tumor cell content and protocol variations including different instruments, reagent lots, and different RNA extraction kits. Variability was evaluated across intratumor biopsies and genomic platforms [RNA sequencing (RNAseq) versus nCounter], and according to protocol variations. RESULTS: Precision analysis of 10 FFPE tumor samples yielded a maximal standard error of 0.94 across HER2DX scores (1-99 scale). High reproducibility of HER2DX scores across 29 FFPE tumors and 20 RNAs between laboratories was evident (correlation coefficients >0.98). The probability of identifying score differences >5 units was ≤5.2%. No significant variability emerged based on platform instruments, reagent lots, RNA extraction kits, or TagSet thaw/freeze cycles. Moreover, HER2DX displayed robustness at low tumor cell content (10%). Intratumor variability across 212 biopsies (106 tumors) was <4.0%. Concordance between HER2DX scores from 30 RNAs on RNAseq and nCounter platforms exceeded 90.0% (Cohen's κ coefficients >0.80). CONCLUSIONS: The HER2DX assay is highly reproducible and robust for the quantification of recurrence risk, pCR likelihood, and ERBB2 mRNA expression in early-stage HER2-positive breast cancer.

Direct detection of bacterial genomic DNA using gold nanoparticle probes.

The molecular probes and associated instrumentation necessary to perform genetic analyses are typically expensive, complex, and prone to error. While techniques such as real-time polymerase chain reaction (PCR) and gene expression analysis have provided a wealth of information previously unattainable, their utility in clinical diagnostics has yet to be realized due to the aforementioned limitations. Nanosphere Inc. has developed a gold nanoparticle-based platform for sequence specific DNA detection that is well-suited for clinical diagnostics due to its cost-effectiveness, simplicity, and accuracy. Thirteen nanometer gold nanoparticle probes, stabilized by a shell of oligonucleotides using proprietary attachment chemistries, enable highly sensitive and specific detection of bacterial genomic DNA sequences without synthetic amplification techniques on a glass array. After silver staining, light scattered by the nanoparticle probes is collected with robust, cost-effective instrumentation. It is the unique features of Nanosphere's detection methodology that provide the necessary steps forward to allow for genetic analyses to become part of routine clinical diagnostics.

Euthanasia of small animals with nitrogen; comparison with intravenous pentobarbital.

Intravenous pentobarbital (with or without addition of saturated potassium chloride) was compared with nitrogen gas exposure for euthanasia of small animals (dogs, cats, and rabbits) in a humane society environment. Initially, electrocardiographic) and electroencephalographic monitoring were used to establish the time of death in presedated animals given either pentobarbital or exposed to nitrogen; later, nitrogen euthanasia alone was studied. Sedation with acepromazine delayed the effects of nitrogen exposure. Addition of intravenous potassium chloride dramatically enhanced the effects of pentobarbital. When sedation was omitted, nitrogen was quantitatively as effective as pentobarbital alone. An adequate flow of nitrogen is essential and the concentration of oxygen in the euthanasia chamber must be monitored.

Measurement of plasma fentanyl concentration: comparison of three methods.

The measurement of plasma fentanyl concentration by radioimmunoassay (RIA) and gas liquid chromatography (GLC) using either a flame ionization detector (FID) or nitrogen/phosphorus detector (NPD) has been compared. RIA is a satisfactory but expensive method of measuring plasma fentanyl concentration. GLC using an FID is not as satisfactory as RIA, but when an NPD is used the results are equal to those of RIA. In addition, other analgesics which are chemically similar to fentanyl, such as alfentanil, may also be measured by the GLC/NPD combination using the same set of operating conditions. By contrast, an RIA method is usually specific for only one compound, and measurement of additional drugs would almost certainly necessitate the development of new assay kits for each one.

Assessing damage to the respiratory tract following thermal injury.

Physicians may institute respiratory therapy at several points along the course of treatment of the burn patient. In the emergency room, they may have to introduce an airway before they can transport the patient to a burn unit. Physicians may have to administer oxygen or place the patient on mechanical ventilation to reverse ventilatory failure, start treatment of the damage to the airways, or prevent or respond to respiratory complications. Any time a physician must choose respiratory therapy, he must base his decision on accurate assessments of the upper and lower airways.