A high-sensitivity lanthanide nanoparticle reporter for mass cytometry: tests on microgels as a proxy for cells.

This paper addresses the question of whether one can use lanthanide nanoparticles (e.g., NaHoF4) to detect surface biomarkers expressed at low levels by mass cytometry. To avoid many of the complications of experiments on live or fixed cells, we carried out proof-of-concept experiments using aqueous microgels with a diameter on the order of 700 nm as a proxy for cells. These microgels were used to test whether nanoparticle (NP) reagents would allow the detection of as few as 100 proteins per "cell" in cell-by-cell assays. Streptavidin (SAv), which served as the model biomarker, was attached to the microgel in two different ways. Covalent coupling to surface carboxyls of the microgel led to large numbers (>10(4)) of proteins per microgel, whereas biotinylation of the microgel followed by exposure to SAv led to much smaller numbers of SAv per microgel. Using mass cytometry, we compared two biotin-containing reagents, which recognized and bound to the SAvs on the microgel. One was a metal chelating polymer (MCP), a biotin end-capped polyaspartamide containing 50 Tb(3+) ions per probe. The other was a biotinylated NaHoF4 NP containing 15 000 Ho atoms per probe. Nonspecific binding was determined with bovine serum albumin (BSA) conjugated microgels. The MCP was effective at detecting and quantifying SAvs on the microgel with covalently bound SAv (20 000 SAvs per microgel) but was unable to give a meaningful signal above that of the BSA-coated microgel for the samples with low levels of SAv. Here the NP reagent gave a signal 2 orders of magnitude stronger than that of the MCP and allowed detection of NPs ranging from 100 to 500 per microgel. Sensitivity was limited by the level of nonspecific adsorption. This proof of concept experiment demonstrates the enhanced sensitivity possible with NP reagents in cell-by-cell assays by mass cytometry.

Synthesis and mass cytometric analysis of lanthanide-encoded polyelectrolyte microgels.

This article describes the synthesis and characterization of two series of functional polyelectrolyte copolymer microgels intended for bioassays based upon mass cytometry, a technique that detects metals by inductively coupled plasma mass spectrometry (ICP-MS). The microgels were loaded with Eu(III) ions, which were then converted in situ to EuF(3) nanoparticles (NPs). Both types of microgels are based upon copolymers of N-isopropylacrylamide (NIPAm) and methacrylic acid (MAA), poly(NIPAm/VCL/MAA) (VCL = N-vinylcaprolactam, V series), and poly(NIPAm/MAA/PEGMA) (PEGMA = poly(ethylene glycol)methacrylate, PG series). Very specific conditions (full neutralization of the MAA groups) were required to confine the EuF(3) NPs to the core of the microgels. We used mass cytometry to measure the number and the particle-to-particle variation of Eu ions per microgel. By controlling the amount of EuCl(3) added to the neutralized microgels. we could vary the atomic content of individual microgels from ca. 10(6) to 10(7) Eu atoms, either in the form of Eu(3+) ions or EuF(3) NPs. Leaching profiles of Eu ions from the hybrid microgels were measured by traditional ICP-MS.

[Electroacupuncture improves cognitive function by inhibiting NF-κB activity in rats with chronic fatigue syndrome].

OBJECTIVE: To observe the effect of electroacupuncture (EA) on the expression of NF-κB p65 in hippocampus and the morphology of hippocampus in rats with chronic fatigue syndrome (CFS), so as to explore its mechanism in improving cognitive dysfunction of CFS. METHODS: Forty-eight SD rats were randomly divided into control, model, EA and inhibitor groups (n=12 in each group). The CFS model was established by multi-factor compound stress stimulation method. Rats of the EA group received EA (50 Hz, 1 mA) at "Baihui" (GV20), Emotional Area I and bilateral Sensory Area for 30 min, once daily for 15 days. For rats in the inhibitor group, pyrrolidine dithiocarbamate (100 mg·kg-1·d-1) was injected intraperitoneally, once a day for 15 days. Learning and memory ability was evaluated by Morris water maze test. HE staining was used to observe the morphology of hippocampus. Western blot was used to determine the expression level of NF-κB p65 in hippocampus. RESULTS: After mode-ling, the general status score was increased （P<0.01）, the escape latency was prolonged(P<0.01), the times of crossing the platform was decreased(P<0.01), and the expression level of NF-κB p65 in hippocampus tissue was significantly increased (P<0.05) in the model group compared with the control group. Compared with the model group, the general status score was decreased （P<0.01）, the escape latency was shortened(P<0.01), the times of crossing the platform was increased(P<0.01), and the expression level of NF-κB p65 in hippocampus tissue was significantly decreased (P<0.05) in the EA and inhibitor groups. HE staining showed that in the model group, the hippocampal nerve cells were arranged disorderly, the structure was loose, and the number of apoptotic bodies and inflammatory cells was significantly increased. The degree of tissue damage of the EA and inhibitor groups was milder than that of the model group. CONCLUSION: EA can improve the cognitive function in CFS rats, which may be associated with its effect in inhibiting the expression of NF-κB and reducing the inflammation response in hippocampus.

[Effect of electroacupuncture on the biochemical indices of bone and bone collagen metabolism and TNF-alpha in osteoporosis model rats without ovaries].

OBJECTIVE: To explore the mechanisms of electroacupuncture (EA) on postmenopausal osteoporosis (PMO). METHODS: Sixty female SD rats aged 6 months were selected, resected double ovaries and fed for 90 days in order to make the model of experimental osteoporosis, and then, they were randomly divided into a model control group (without any therapy), a routine acupoints applying group, which were treated with EA at "Pishu" (BL 20), "Weishu" (BL 21), "Shenshu" (BL 23) and "Qihaishu" (BL 24), and an EA with tonification method (EAT) group at "Guanyuan" (CV 4) and "Zusanli" (ST 36), and an EA with dispersing blood stasis method (EAD) group at "Shenshu" (BL 23), "Geshu" (BL 17) and "Dazhu" (BL 11), and an EA with tonification and dispersing blood stasis method (EATD) group at "Guanyuan" (CV 4), "Zusanli" (ST 36), "Shenshu" (BL 23), "Geshu" (BL 17) and "Dazhu" (BL 11), 12 rats in each group. EA therapy was performed once a day, 30 min each time. Then these rats rested for 1 day after consecutive treatment for 6 days. The treatment lasted for 12 weeks. Enzyme linked immunosorbent assay (ELISA) was used to examine the level of TNF-alpha, and the level of E2, osteocalin (BGP) and carboxyterminal propeptide of type I procollagen (PICP), carboxyterminal telopeptide of type I lollagen (ICTP) by radioimmunoassay, and alkaline phosphatase by p-nitrophenylphosphate method, and Ca, P, Deoxypyridinoline (DPD) and creatinine (Cr) in urine were detected by automatic biochemistry analyzer. RESULTS: Compared with model control group, the level of DPD/Cr, Ca/Cr, and the serum contents of ALP, BGP, TNF-alpha and ICTP in all EA groups decreased significantly (all P < 0.01), the level of E2 and PICP in serum increased significantly (all P < 0.01). Compared with EATD group, the level of DPD/Cr and Ca/Cr in the other three EA groups had no statistical differences (all P > 0.05), but the level of ALP, BGP, TNF-alpha and ICTP in serum increased significantly (all P < 0.01), the level of E2 and PICP in serum decreased significantly (all P < 0.01). CONCLUSION: Acupuncture can reduce significantly the level of DPD/Cr, Ca/Cr, and the serum contents of ALP, BGP, TNF-alpha and ICTP, increase significantly the level of E2 and PICP in PMO model rats and EA with tonification and dispersing blood stasis method is superior significantly to the other routine acupuncture methods.

Highly luminescent lead sulfide nanocrystals in organic solvents and water through ligand exchange with poly(acrylic acid).

Hydrophobic lead sulfide quantum dots (PbS/OA) synthesized in the presence of oleic acid were transferred from nonpolar organic solvents to polar solvents such as alcohols and water by a simple ligand exchange with poly(acrylic acid) (PAA). Ligand exchange took place rapidly at room temperature When a colloidal solution of PbS/OA in tetrahydrofuran (THF) was treated with excess PAA, the PbS/PAA nanocrystals that formed were insoluble in hexane and toluene but could be dissolved in methanol or water, where they formed colloidal solutions that were stable for months. Ligand exchange was accompanied by a small blue shift in the band-edge absorption, consistent with a small reduction in particle size. While there was a decrease in quantum yield associated with ligand exchange and transfer to polar solvents, as is commonly found for colloidal quantum dots, the quantum yields determined were impressively high: PbS/OA in toluene (82%) and in THF (58%); PbS/PAA in THF (42%) and in water (24%). The quantum yields for the PbS/PAA solutions decreased over time as the solutions were allowed to age in the presence of air.