The MGT substrate with high reproducibility, sensitivity, and selectivity was then developed as an immunoassay SERS probe (MGT-Abs-CuPc) through the immobilization of the PD-L1 Abs and CuPc for PD-L1 analysis on TNBC surfaces (Fig. 5A). As shown in fig. S5A, upon the introduction of CuPc molecules to MGT, the UV-vis spectrum exhibits two peaks at 620 and 690 nm, which correspond to the adsorption of CuPc. After the immobilization of Abs to MGT, we observed an additional peak at ~260 nm, which should be attributed to adsorption of Abs. All these three additional peaks show the successful fabrication of the MGT-Abs-CuPc nanoprobe. To further confirm Abs conjugation to the nanostructures, we performed SDS–polyacrylamide gel electrophoresis (PAGE) imaging of MGT, Abs, and Abs-modified MGT (MGT-Abs). SDS-PAGE is a powerful analytical technique for protein identification in complex biological samples and has been widely used in biological studies for decades. As shown in fig. S5B, MGT-Abs [fig. S5B (III)] was higher than both MGT [fig. S5B (I)] and Abs [fig. S5B (II)], indicating the different molecular weights: MGT-Abs > MGT > Abs. SDS-PAGE imaging in fig. S5B (I to III) definitively showed the successful conjugation between the nanostructure and Abs. In addition, we also carried out FTIR spectroscopy (fig. S5C) to investigate the conjugation. Compared to MGT, MGT-Abs showed an additional infrared band assigned to the C–N stretching vibration at 1400 cm−1 and the C=O stretching vibration at 1620 cm−1. These two additional peaks were attributed to the formation of an amide bond at the interface between MGT and Abs. Both SDS-PAGE images and FTIR results revealed that Abs has been successfully conjugated to the nanostructures. We carried out confocal imaging to verify the recognition ability of MGT-Abs to the PD-L1 protein. We conducted three parallel experiments, which are shown in fig. S5 (D to G). No fluorescence signal was obtained after the coincubation of MGT and Cy5-labeled PD-L1. Moreover, before incubating with Cy5-labeled PD-L1, MGT-Abs also showed weak fluorescence. However, after the incubation with Cy5-labeled PD-L1, confocal images exhibited a bright fluorescence signal, indicating the high affinity and recognition ability of MGT-Abs to the PD-L1 protein. We incubated the developed probe with high SERS activity with TNBCs to quantify the PD-L1 expression on the surface of different TNBCs (Fig. 5A) using Raman spectroscopy. Temperature and incubation time are two important factors that determine the overall SERS signals. As shown in fig. S5 (H and I), the maximum SERS response was achieved at 37°C and 60 min. The amount of CuPc and Abs immobilized on MGT was another important experimental parameter. As the concentration of CuPc and Abs progressively increased, we observed the Raman peak at 1530 cm−1 to first increase and then decrease after the concentration reached 10 mM for CuPc and 100 μM for Abs, indicating that the optimal concentration of CuPc is 10 mM, while that of Abs is 100 μM (fig. S5, J and K). Moreover, we examined the cytotoxicity of the MGT-Abs-CuPc probe with a 3-(4,5-dimethylthiazol-2-yl)-2-diphenyltetrazolium bromide assay. As a result, the variability of HCC38, MDA-MB-231, and MCF-7 cells remained approximately 98% (15 μM, 120 min; fig. S6), indicating the low cytotoxicity of the probe. After optimizing experimental conditions and assessing biocompatibility, we conducted three parallel experiments to evaluate the PD-L1 expression levels on three different types of TNBC (HCC38, MCF-7, and MDA-MB-231) surfaces. HCC38 cells were first used as model cells. As shown in Fig. 5B, Raman intensity increased with an increasing number of cells captured on the silicon foil. The intensity of the Raman peak at 1530 cm−1 was used to reflect the amount of the MGT-Abs-CuPc probe attached to the cell surface. The intensity of the Raman peak (I) was proportional to the logarithmic value of captured cancer cells (N) in the range from 5 × 102 to 5 × 105 cells (R = 0.990, n = 7; Fig. 5C), which was consistent with the kinetics mechanism of the antigen-Abs reaction in the Abs excess zone (fig. S7A). The linear regression is shown in Eq. 1

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Impressive, but I  wish that he’d given the money to programs that help students at non-elite universities rather than already rich ones such as Hopkins. That would do wider good for the country.

Inventor(s): Brian Gventer (Fort Worth, TX), Joel McCarty (Oklahoma City, OK), Ken Nguyen (Fort Worth, TX), Kevin Asbury (Van Alstyne, TX), Mike Lowry (Fort Worth, TX) Assignee(s): FedEx Supply Chain Logistics Electronics, Inc. (Fort Worth, TX) Law Firm: McGuire Woods LLP (Local + 9 other metros) Application No., Date, Speed: 14075590 on 11/08/2013 (1859 days app to issue)

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Current genetic and archeological evidence allows for inland, coastal, or multiple pathways to peopling of the Americas.

Funeral services were conducted Friday afternoon, April 27, 2018 at Hutcheson’s Memorial Chapel with Rev. Randy Rooks officiating.

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Mrs. Helen Mills, age 88, of Cedartown, passed away on Tuesday April 24, 2018 after an extended illness.

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