Pcr tube

The reference spectral responses and effective mass attenuation coefficients needed for MD were obtained by scanning a calibration phantom. A customized PMMA (polymethyl methacrylate) phantom held 200 µL Eppendorf PCR tubes containing AuNP solutions of various concentrations (0.5, 1, 2, 4, 8 and 10 mg/mL), hydroxyapatite (HA) rods (HA 50, 100, 200, 400 and 800 mg/cm³; HA acquired from QRM GmbH), water, and lipid (Figure 1). To evaluate the effect of soft tissue on the material attenuation profile, a layer of lamb red meat was wrapped around the phantom. The calibration phantoms were used to evaluate the linearity of the response to AuNP and HA concentrations and evaluate the spectroscopic response of the detector for different materials.
The same scanning protocol was used to image a material calibration phantom (Figure 1) to provide reference spectral responses and effective mass attenuation coefficients. The reconstructed spectral response images of the phantom were analysed [38 ]. Linear regression analysis determined the relationship between spectral response and material concentration (linearity response) [39 ]. Furthermore, the relationship between the spectral response and the energy thresholds was evaluated graphically (spectral response).

SAFS and SASL had been captured in 2011 and 2014–2018 for health evaluation at PSJ, Peru (15°22’S, 75°12’W) under Peruvian permits RJ 23–2011, 024–2014, 229–2015, and 019-2016-SERNANP-RNSIIPG. All sample collection was conducted in November, except for 2017 (February) and 2018 (April). Adult animals were anesthetized under the supervision of a veterinarian. Pups under 6 months of age were either manually restrained or anesthetized based on protocol needs for concurrent projects. A sterile cotton-tipped applicator (Cardinal Health sterile cotton tipped applicator with plastic shaft, Cardinal Health, Dublin, Ohio) was used to collect urogenital swabs (URO) from the prepuce or vulva, conjunctival swabs (CON) from the inner palpebrae, and oropharyngeal swabs (ORO) from the dorsal aspect of the oral cavity along the soft palate. All swabs were placed without media in microcentrifuge (Eppendorf PCR tubes, Eppendorf North America, Hauppauge, NY) tubes and promptly placed on ice. Samples were frozen within 12 hours and were maintained at -80°C until DNA extraction.

DNA dissolved in TE buffer (Thermo Fisher no. 12090015, Invitrogen, Waltham, MA, USA) was brought to room temperature slowly from −20 °C. Using universal primers (ITS1 and ITS4), the fungal ITS gene was amplified. First, the solution (500 µL) was added to PCR tubes (Eppendorf no. 0030124332, Hamburg, Germany). Then, 0.5 pmol of ITS1 and ITS4 was added following by deoxynucleotide triphosphates (dNTPs, 200 μM each) (Promega no. U1511, Madison, OH, USA), 0.5 U DNA polymerase (Promega no. D1501, Madison, OH, USA), supplied PCR buffer (ThermoFisher no. 14966123, Platinum II Green PCR Buffer) and water. PCR was performed as follow: 1 cycle (98 °C for 2 min) for initial denaturation; 25 cycles (98 °C for 15 secs; 60 °C for 30 secs; 72 °C for 30 sec) for annealing and extension, and 1 cycle (72 °C for 10 min) for final extension of the amplified DNA (Eppendorf Mastercycler gradient, Hamburg, Germany) [20] (link).