Label-Free and Real-Time Optical Detection of Affinity Binding of the Antibody on Adherent Live Cells.

Oblique-incidence reflectivity difference (OIRD) is a novel real-time, label-free, and nondestructive optical detection method and exhibits encouraging application in the detection of antibody/DNA microarrays. In this study, for the first time, an OIRD label-free immunoassay was achieved by using adherent live cells as the probe. The cells were cultured on glass cells, and the affinity binding of antibodies targeted on the HLA class I antigen of the cell surface was detected with an OIRD. The results show that an OIRD is able to detect the binding process of anti-human HLA-A, B, and C antibodies on MDA-MB-231 cells and HUVEC cells. Control experiments and complementary fluorescence analysis confirmed the high detection specificity and good quantitative virtue of the OIRD label-free immunoassay. Label-free OIRD imaging analysis of cell microarrays was further demonstrated successfully, and the underlying optical mechanism was revealed by combining the theoretical modeling. This work explores the use of live cells as probes for an OIRD immunoassay, thus expanding the potential applications of the OIRD in the field of pathological analysis, disease diagnosis, and drug screening, among others.

Co3O4 Nanoparticles Uniformly Dispersed in Rational Porous Carbon Nano-Boxes for Significantly Enhanced Electrocatalytic Detection of H2O2 Released from Living Cells.

A facile and ingenious method to chemical etching-coordinating a metal-organic framework (MOF) followed by an annealing treatment was proposed to prepare Co3O4 nanoparticles uniformly dispersed in rational porous carbon nano-boxes (Co3O4@CNBs), which was further used to detect H2O2 released from living cells. The Co3O4@CNBs H2O2 sensor delivers much higher sensitivity than non-etching/coordinating Co3O4, offering a limit of detection of 2.32 nM. The wide working range covers 10 nM-359 µM H2O2, while possessing good selectivity and excellent reproducibility. Moreover, this biosensor was used to successfully real-time detect H2O2 released from living cells, including both healthy and tumor cells. The excellent performance holds great promise for Co3O4@CNBs's applications in electrochemical biomimetic sensing, particularly real-time monitor H2O2 released from living cells.

[Methylation in Promoter Region of SLC6A2 Gene in Heart Failure Patients and Its Correlation with Qi Deficiency/Blood Stasis Syndrome].

OBJECTIVE: To explore the methylation status in promoter region of norepinephrine transporter gene (NET, SLC6A2) in heart failure ( HF) patients and its correlation with qi deficiency/blood stasis syndrome (QDS/BSS). METHODS: Thirty-six patients with heart failure (NYHA classification III to IV) were recruited in the study (as the heart failure group) and their scores of QDS/BSS were evaluated. Besides, a healthy elderly group (30 cases) and a healthy youth group (30 cases) were also set up. They were recruited from Physical Examination Center of Fujian Provincial Hospital. Pyrosequencing was applied to detect the methylation in promoter region of SLC6A2 gene, and the total methylation index (MTI) of CpG island was calculated. The correlation between the methylation status in promoter region of SLC6A2 and scores of QDS/BSS was assessed using Pearson and Partial analyses. Risk factors were screened and adjusted using Logistic regression. RESULTS: By one-factor analysis of variance, the total MTI in the HF group (219.72% ± 54.03%) was obviously higher than that in the healthy elderly group (194.47% ± 34.92%) and the healthy youth group (161.60% ± 41.11%) (all P < 0.05). Meanwhile, the total MTI was higher in the healthy elderly group than in the healthy youth group (P < 0.01). By covariance analysis , after controlling age and BMI, the total MTI was higher in the HF group than in the healthy elderly group (P = 0.041), while it was higher in the healthy elderly group than in the healthy youth group (P = 0.016). Age was found to play an essential role in affecting MTI of SLC6A2 gene promoter region among the 3 groups (F = 16.447, P = 0.01). The total MTI was quite lower in the healthy youth group. Results of Partial correlation analysis showed MTI was positively correlated with scores of qi deficiency and blood stasis respectively (r = 0.494 and 0.419 respectively, both P < 0.05). Logistic regression analysis showed after adjusting confounding factors, the relative risk (OR value) of total MTI of SLC6A2 gene in promoter region was 1.038 (95% CI, 1.006 to 1.071, P = 0.020). CONCLUSIONS: Abnormally elevated methylation of the promoter region of SLC6A2 gene is one of risk factors for HF. In addition, the degree of methylation of the promoter region of SLC6A2 gene was positively correlated with the severity of QDS/BSS.

Cell repelling agar@paper interface assisted probing of the tumor spheroids infiltrating natural killer cells.

Scaffold-based culture is one of the effective methods to resemble three-dimensional (3D) cells model in vitro. An agar@lens paper hybrid scaffold was prepared by one-pot dip-coating. The lens paper's cellulose fiber networks are the scaffold's backbone. The agar gel seized the gaps between the fibrous structures that can improve the paper scaffold's optical transparency and prevent cells from spreading on the scaffold. The SEM and light microscope images showed that the agar gel on the bottom of the paper and the cellulose fiber of the paper formed micro-well structures. Without staining, the cells growing on the agar@paper scaffold can be directly observed under a light microscope. Cells aggregated between the cellulose fibers and formed spheroids within 24 h. The cell spheroids can be non-enzymatically retrieved from the agar@paper scaffold because of the cell-repelling property of agar. The agar@paper scaffold was applied for co-culturing tumor cells (MDA-MB-231, DU 145) and natural killer cells (NKs, NK-92). Using the agar@paper scaffolds, the tumor-infiltrating NKs can be separated from floating NKs that did not attack the tumor spheroids. The effect of NKs infiltrating on tumor spheroids size was characterized. The results showed that NKs attacking the spheroids grown on agar@paper scaffold can be readily tracked because of the improved optical transparency. Higher NKs: tumor cells ratio resulted in a high percentage of tumor infiltrating NKs. The separated NKs can be further tested to reveal their biological characteristics. Both agar and lens paper is accessible for most biological labs, highlighting the potential of agar@paper scaffold in 3D culture.

Discovery of polypyridyl iridium(III) complexes as potent agents against resistant Candida albicans.

The increasing emergence and spread of drug resistant Candida albicans represent a serious challenge for effective treatment and call for the development of new therapeutic options. To address this need, we synthesized a series of polypyridyl iridium(III) complexes and studied their antimicrobial activities. Herein, one lead iridium(III) complex Ir-3 [(ptpy)2Ir(dppz)]PF6, with ptpy = 2-(p-tolyl)pyridine and dppz = dipyrido[3,2-a:2',3'-c]phenazine, exhibited potent and broad-spectrum antifungal activities against drug-resistant pathogens and low hemolytic activity toward mammalian cells. Furthermore, Ir-3 showed low tendency to induce resistance, displayed biofilm inhibition and eradication activities. Significantly, Ir-3 exhibited potent in vivo antifungal activity in a murine model of disseminated candidiasis. This study may pave the way for the development of novel antifungal agent based upon polypyridyl iridium(III) complex to combat antifungal resistance.

[The role of peroxisome proliferator-activated receptor gamma in proliferation of cardiac non myocytes].

AIM: To investigate the effect of PPARgamma activators on inhibition of cardiac non myocytes (CNM) proliferation and the PPARgamma-dependent pathway possibly involved. METHODS: Angiotensin II was used to induce proliferation of primarily cultured CNM from neonatal rats, and pioglitazone or 15-deoxy-delta12,14-prostaglandin J2 (15d-PGJ2) was applied to these CNM in various dosages in vitro. MTT assay and 3H-TdR uptake were determined to estimate proliferation of CNM, and transient transfection of reporter gene containing PPRE from ACO promoter (PPRE-pGL3) with or without PPARgamma expression plasmid (PPARgamma-pSG5) to CNM was performed to determine the control of target-gene transcription. RESULTS: Angiotensin II caused a significant increase in MTT value and 3H-TdR uptake in CNM, which could be significantly reversed by pioglitazone and 15d-PGJ2 in a dose-dependent manner. Transient cotransfection of PPRE-pGL3 with PPARgamma-pSG5 to CNM resulted in significant increase in luciferase activity compared with that without PPARgamma-pSG5 cotransfection. Pioglitazone and 15d-PGJ2 induced increase in luciferase activity also in a dose-dependent manner. CONCLUSION: Pioglitazone and 15d-PGJ2, as the activators of PPARgamma, inhibit proliferation of CNM from neonatal rats, the effect may be related to the activation of PPARgamma.