Assessment of Preanalytical Cerebrospinal Fluid Handling and Storage Factors on Measurement of Aß1-42, Aß1-40, and pTau181 Using an Automated Chemiluminescent Platform.

BACKGROUND: Standardizing cerebrospinal fluid (CSF) laboratory protocols will improve the reliability and availability of clinical biomarker testing required for prescription of novel Alzheimer disease (AD) therapies. This study evaluated several preanalytical handling and storage factors common to ß-amyloid1-42 (Aß1-42), ß-amyloid1-40 (Aß1-40), and phosphorylated tau (pTau181) concentrations including storage at different temperatures, extended cap contact, various mixing methods, and multiple freeze-thaw cycles. METHODS: Aß1-42, Aß1-40, and pTau181 concentrations were measured using LUMIPULSE G1200 automated assays. Samples were collected in polypropylene tubes of various volumes. Sample cap-contact was evaluated by storing samples in upright and inverted positions at either 4°C for 1 week or -80°C for 1 month. To assess mixing methods, samples were freeze-thawed and mixed by inversion, vortex, horizontal roller, or unmixed prior to assay sampling. The impact of successive freeze-thaw cycles was assessed through freezing, thawing, and analyzing CSF samples. RESULTS: Short-term storage at 4°C did not affect Aß1-42, Aß1-40, or pTau181 measurements in any tube type. Tube cap contact affected Aß1-42 in 2.5 mL tubes and pTau181 levels in 10 mL tubes. No difference was observed between mixing methods. After 4 freeze-thaw cycles, Aß1-42 significantly decreased but Aß1-40 remained unchanged. Utilizing the Aß1-42/Aß1-40 ratio, Aß1-42 values normalized, maintaining ratio values within ±5% of baseline measurements. CONCLUSIONS: Storage of CSF at 4°C for 1 week or -80°C for 1 month did not significantly affect Aß1-42, Aß1-40, pTau181, or associated ratio measurements. Tube cap-contact impacted pTau181 and pTau181/Aß1-42 values in larger tubes. Mixing methods are equivalent. The Aß1-42/Aß1-40 ratio compensates for freeze-thaw variability up to 4 cycles.

Effect of Patient-Specific Preanalytic Variables on CSF Aß1-42 Concentrations Measured on an Automated Chemiluminescent Platform.

BACKGROUND: Cerebrospinal fluid (CSF) biomarkers are increasingly used to confirm the accuracy of a clinical diagnosis of mild cognitive impairment or dementia due to Alzheimer disease (AD). Recent evidence suggests that fully automated assays reduce the impact of some preanalytical factors on the variability of these measures. This study evaluated the effect of several preanalytical variables common in clinical settings on the variability of CSF ß-amyloid 1-42 (Aß1-42) concentrations. METHODS: Aß1-42 concentrations were measured using the LUMIPULSE G1200 from both freshly collected and frozen CSF samples. Preanalytic variables examined were: (1) patient fasting prior to CSF collection, (2) blood contamination of specimens, and (3) aliquoting specimens sequentially over the course of collection (i.e., CSF gradients). RESULTS: Patient fasting did not significantly affect CSF Aß1-42 levels. While assessing gradient effects, Aß1-42 concentrations remained stable within the first 5 1-mL aliquots. However, there is evidence of a gradient effect toward higher concentrations over successive aliquots. Aß1-42 levels were stable when fresh CSF samples were spiked with up to 2.5% of blood. However, in frozen CSF samples, even 0.25% blood contamination significantly decreased Aß1-42 concentrations. CONCLUSIONS: The preanalytical variables examined here do not have significant effects on Aß1-42 concentrations if fresh samples are processed within 2 h. However, a gradient effect can be observed on Aß1-42 concentrations after the first 5 mL of collection and blood contamination has a significant impact on Aß1-42 concentrations once specimens have been frozen.

The use of focused ultrasound for the treatment of cutaneous allodynia associated with chronic migraine.

Chronic migraines (CM) are the third most common disease and are refractory to medical treatment in 15% of patients. Currently, temporary relief is achieved with steroid blocks or pulsed radiofrequency ablation, which have short-term benefits. Our project aims to develop a non-invasive treatment for medically refractory chronic migraine, which does not require a permanent implant. This project investigates the safety and effectiveness of pulsed focused ultrasound (FUS) in a validated rodent headache model of cutaneous allodynia associated with chronic migraine (CM) as compared to sumatriptan and ablative lesioning. We demonstrate a significant reduction in mechanical thresholds as measured through Von Frey filaments in CM in the forepaw and periorbital region (p < 0.001). Sumatriptan and pulsed FUS both significantly improve thresholds at day 3 after treatment in the periorbital region. Ablative lesioning has no effect. This study provides initial evidence that FUS may provide an important therapeutic option for patients suffering from CM.