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Two-dimensional (2D) nanosheets as carriers have shown promising potential for surface-displaying or loading various drugs. Nevertheless, developing sheet-like materials themselves into an immunoregulator has never been realized so far. In this study, we take advantage of the immunoregulatory effects of rare earth elements themselves and develop water-soluble erbium–dysprosium 2D nanosheets (2D NSs). Such 2D NSs can target lymph nodes and activate macrophages to improve vaccine efficacy in mice significantly. Transcriptome analysis further reveals that six critical molecules (Msr1, Ccr2, Serpinb9, Klrk1, Klrd1, Klrc1) closely correlate with 2D NS-mediated immunoregulation in vivo. For the first time, the present work realizes a proof-of-concept for designing immunoregulatory 2D NSs and shows a promising potential of 2D NSs for improving the immunoprophylaxis/immunotherapy of vaccines., 2D NSs target lymph nodes and activate macrophages to enhance vaccine-induced immune responses via regulating six critical genes (Ccr2, Serpinb9, Klrk1, Klrd1, Klrc1, Msr1).

Introduction Erythropoiesis slowly decreases with increasing age, which may be reflected in red blood cell (RBC) parameters. This multicentre collaborative study aimed to investigate the changes in erythropoiesis with increasing age in a healthy Chinese population. Methods A total of 14,591 healthy individuals (6,713 aged at least 60 y and 7,878 aged below 60 y) from seven cities across China were enrolled. K2-EDTA anticoagulant blood samples were analysed. The results are presented as median and 2.5-97.5th percentile. Results RBC parameters showed some differences between the two groups divided by the age of 60 in the Chinese population. The median, 2.5th and 97.5th percentile values of RBC, haemoglobin (HGB) and haematocrit (HCT) in patients aged ≥ 60 y were significantly lower than in those ˂ 60 y. The values of mean cell volume (MCV), mean cell haemoglobin (MCH) and red cell distribution width (RDW) were higher in the group aged ≥ 60 y. Men had significantly higher RBC, HGB, HCT, MCV, MCH and RDW indices than women. The prevalence of anaemia gradually increased with age in men and was higher than that in women after 50. The median haemoglobin and MCV in Nanning and Guangzhou were lower than those in other regions. Conclusion RBC parameters varied with increasing age and differed between males and females, indicating that erythropoiesis decreases in the elderly Chinese population. Subsequent studies should be conducted for age- and sex-specific reference intervals in healthy elderly Chinese populations.

Intermolecular forces constrain peptide conformation. However, the role of each intermolecular force in constraining peptide conformation remains poorly understood. In this work, we show that aromatic-aromatic interactions drive peptides into β-sheets, and the hydrophobic effect determines the assembly speed of peptides. By using intermolecular forces to artificially control the assembly of β-sheets, a multi-modal analytical system was developed that allows five readouts and dual qualitative-quantitative analysis, and satisfies both point-of-care testing (POCT) and laboratory-based testing. For Mycoplasma Pneumoniae diagnosis, this system eradicates misdiagnosis (from 30 % to 0 %) and broadens the linear range by three-fold, both of which are critical for guiding therapy. This work not only illustrates exact roles of intermolecular forces in driving the formation of β-sheets, but also provides a guideline for the construction of a multi-modal analytical system for disease diagnosis.

Molecular profiling of circulating extracellular vesicles (EVs) provides a promising noninvasive means to diagnose, monitor, and predict the course of metastatic breast cancer (MBC). However, the analysis of EV protein markers has been confounded by the presence of soluble protein counterparts in peripheral blood. Here we use a rapid, sensitive, and low-cost thermophoretic aptasensor (TAS) to profile cancer-associated protein profiles of plasma EVs without the interference of soluble proteins. We show that the EV signature (a weighted sum of eight EV protein markers) has a high accuracy (91.1 %) for discrimination of MBC, non-metastatic breast cancer (NMBC), and healthy donors (HD). For MBC patients undergoing therapies, the EV signature can accurately monitor the treatment response across the training, validation, and prospective cohorts, and serve as an independent prognostic factor for progression free survival in MBC patients. Together, this work highlights the potential clinical utility of EVs in management of MBC., A thermophoretic aptasensor can be used to profile cancer-associated proteins of extracellular vesicles (EVs) in patients’ plasma. Here, the authors use this technique to develop an EV-signature able to discriminate metastatic breast cancer, monitor treatment response, and predict patients’ progression-free survival.

Our recent publication illustrates the critical role of phenylalanine-mediated aromatic-aromatic interactions in determining the assembly of peptidic β-sheets. However, the effect of phenylalanine number on regulating the assembly efficacy of peptidic β-sheets remains poorly understood. We herein evaluate the assembly efficacy of β-sheets of a series of oligopeptides which contain 0, 1, 2, or 3 phenylalanine in their molecular backbones. In our assembly system, two phenylalanine (2F) is the minimum number for driving the assembly of β-sheets of oligopeptides. Oligopeptides with three phenylalanine (3F) show significantly increased assembly efficacy of β-sheets compared to that with 2F. These results suggest a positive correlation between the phenylalanine number and assembly efficacy of β-sheets. By improving the assembly efficacy of β-sheets, we further develop a highly sensitive HIV analytical system in which the specific binding of β-sheets with Congo Red induces enhanced fluorescence. For HIV p24 detection, the 3F-based analytical system (0.61 pg/mL) shows a significantly lower limit of detection (LOD) than the 2F-based analytical system (2.44 pg/mL), both of which are more sensitive than commercial ELISA (5 pg/mL) used in the clinic. This work not only illustrates the effect of phenylalanine number on regulating the assembly efficacy of β-sheets but also provides a guideline for the construction of a highly sensitive analytical system of disease diagnosis.

How to balance the sensitivity and signal-to-noise ratio of immunosensor remains many challenges during various diseases diagnosis. Here we develop a new microfluidic immunosensor based on surface-modified mesoporous nanofibers, and simultaneously realize an ultra-sensitivity and high signal-to-noise ratio for the detection of multiple biomarkers. In the current study, we fabricated titanium dioxide (TiO2)-based mesoporous electrospinning nanofibers, and modified nanofiber surface with both octadecylphosphonic acid (OPA) and poly(ethylene oxide)-poly(propylene oxide) triblock copolymer (PEO-PPO-PEO). Such nanofibers as solid substrate are covered on microfluidic channels. The porosity of our nanofibers dramatically increased the adsorption capability of antibodies, realizing an ultra sensitivity of biomarker detection. PEO-PPO-PEO modification can significantly block non-specific absorptions, obtaining a satisfied signal-to-noise ratio. For the detection of HIV p24 and interleukin 5 (IL-5), our immunosensor increased 6.41 and 6.93 fold in sensitivity and improved 504.66% and 512.80% in signal-to-noise ratio, in compared with gold standard immunoassay (ELISA) used in the clinic. Our immunosensor also broaden the linear range for the detection of HIV p24 (0.86–800 pg/ml) and IL-5 (0.70–800 pg/ml), in compared with ELISA which is 5.54–500 pg/ml for HIV p24 and 4.84–500 pg/ml for IL-5. Our work provided a guideline for the construction of advanced point-of-care immunosensor with an ultra-sensitivity and high signal-to-noise ratio for disease diagnosis.

Objective: To evaluate the diagnostic value of fluorescence in situ hybridization (FISH) in bladder cancer. Methods: We enrolled healthy volunteers and patients who were clinically suspected to have bladder cancer and conducted FISH tests and cytology examinations from August 2007 to December 2008. Receiver operating characteristic (ROC) curve analysis was performed and the area under curve (AUC) values were calculated for both the FISH and urine cytology tests. Results: A cohort of 988 healthy volunteers was enrolled to establish a reference range for the normal population. A total of 4807 patients with hematuria were prospectively, randomly enrolled for the simultaneous analysis of urine cytology, FISH testing, and a final diagnosis as determined by the pathologic findings of a biopsy or a surgically-excised specimen. Overall, the sensitivity of FISH in detecting transitional-cell carcinoma was 82.7%, while that of cytology was 33.4% (p

The microfluidic passive control of microparticles largely relies on the hydrodynamic effects of the carrier media such as Newtonian fluids and viscoelastic fluids. Yet the viscoelastic/Newtonian interfacial effect has been scarcely investigated, especially for high-resolution particle separation. Here we report a microfluidic co-flow of Newtonian (water or PBS) and viscoelastic fluids (PEO) for the size-dependent separation of microparticles. The co-flow condition generates a stable viscoelastic/Newtonian interface, giving rise to the wall-directed elastic lift forces that compete with the center-directed lift forces, and efficiently hinders the migration of microparticles from the Newtonian to the viscoelastic fluid in a size-dependent manner. An almost complete separation of a binary mixture of 1 μm and 2 μm polystyrene particles is achieved by the co-flow of water and a very dilute PEO solution (100 ppm), whereas the sole use of water or PEO could not lead to an efficient separation. This co-flow microfluidic system is also applied for the separation of Staphylococcus aureus (1 μm) from platelets (2–3 μm) with >90% efficiencies and purities.

Although conventional enzyme-linked immunosorbent assays (ELISA) and related assays have been widely applied for the diagnosis of diseases, many of them suffer from large error variance for monitoring the concentration of targets over time, and insufficient limit of detection (LOD) for assaying dilute targets. We herein report a readout mode of ELISA based on the binding between peptidic β-sheet structure and Congo Red. The formation of peptidic β-sheet structure is triggered by alkaline phosphatase (ALP). For the detection of P-Selectin which is a crucial indicator for evaluating thrombus diseases in clinic, the ‘β-sheet and Congo Red’ mode significantly decreases both the error variance and the LOD (from 9.7 ng/ml to 1.1 ng/ml) of detection, compared with commercial ELISA (an existing gold-standard method for detecting P-Selectin in clinic). Considering the wide range of ALP-based antibodies for immunoassays, such novel method could be applicable to the analysis of many types of targets.

Non-invasive assays for early cancer screening are hampered by challenges in the isolation and profiling of circulating biomarkers. Here, we show that surface proteins from serum extracellular vesicles labelled with a panel of seven fluorescent aptamers can be profiled, via thermophoretic enrichment and linear discriminant analysis, for cancer detection and classification. In a cohort of 102 patients, including 6 cancer types at stages I–IV, the assay detected stage I cancers with 95% sensitivity (95% confidence interval (CI): 74–100%) and 100% specificity (95% CI: 80–100%), and classified the cancer type with an overall accuracy of 68% (95% CI: 59–77%). For patients who underwent prostate biopsies, the assay was superior to the analysis of prostate-specific antigen levels (area under the curve: 0.94 versus 0.68; 33 patients) for the discrimination of prostate cancer and benign prostate enlargement, and also in the assessment of biochemical cancer recurrence after radical prostatectomy. The assay is inexpensive, fast, and requires small serum volumes (

Platelet function has been described by many laboratory assays, and PL-11 is a new point-of-care platelet function analyzer based on platelet count drop method, which counts platelet before and after the addition of agonists in the citrated whole blood samples. The present study sought to compare PL-11 with other three major more established assays, light transmission aggregometry (LTA), VerifyNow™ aspirin system and thromboelastography (TEG), for monitoring the short-term aspirin responses in healthy individuals. Ten healthy young men took 100 mg/d aspirin for 3-day treatment. Platelet function was measured via PL-11, LTA, VerifyNow and TEG, respectively. The blood samples were collected at baseline, 2 hour, 1 day during the aspirin treatment and 1 day, 5 ± 1 days, 8 ± 1 days after the aspirin withdrawal. Moreover, 90 additional healthy subjects were recruited to establish a reference range for PL-11. Platelet function of healthy subjects decreased significantly 2 hours after 100 mg/d aspirin intake and began to recover during 4-6 days after the aspirin withdrawal. Correlations between methods were PL-11 vs. LTA (r = 0.614, p 0.01); PL-11 vs. VerifyNow (r = 0.829, p 0.01); PL-11 vs. TEG (r = 0.697, p 0.001). There was no significant bias between PL-11 and LTA at baseline (bias = 1.94%, p = 0.804) using Bland-Altman analysis, while the data of PL-11 were significantly higher than LTA (bias = 24.02%, p 0.001) during the aspirin therapy. The reference range for PL-11 in healthy young individuals was from 66.8 to 90.5% (95%CI). When aspirin low-responsiveness was defined as LTA 20%, the cut-off values for each method were, respectively: PL-11 50%, VerifyNow 533 ARU, TEG 60.2%. The results of different platelet function assays were uninterchangeable for monitoring aspirin response and correlations among them were also varied. Correlations among PL-11 and other three major assays suggested the ability of PL-11 to assess the treatment effects of aspirin. But a large cohort study is needed to confirm the cut-off value of aspirin response detected by PL-11.

To investigate the clinical value of new kinds of urinary erythrocyte morphology parameter in discriminating different pathology types of glomerulonephritis.All of the 52 urine samples were from glomerulonephritis patients who had been diagnosed by renal biopsy results. The change of the percentage of acanthocytes, the size of RBC, the shape of RBC between the primary glomerulonephritis (39 cases) and secondary glomerulonephritis (13 cases) urine were detected by AVE-764 fully automatic urine cell analyzer.Acanthocytes could be found in both primary glomerulonephritis and secondary glomerulonephritis. Of the patients whose acanthocytes percentages above 10%, 94.1% had primary glomerulonephritis and 5.9% had secondary glomerulonephritis. The picture of size-shape phase were classified as strip-type, inverted triangle-type and hanging tail-type. 95.2% Strip-type cases were from primary glomerulonephritis patients. Triangle-typenormally cases were all from primary glomerulonephritis patients. Hanging tail-type cases were all from secondary glomerulonephritis.High acanthocytes percentage is most common in primary glomerulonephritis, going with the size and shape of RBC can be useful in the differential diagnosis of different pathology types of glomerulonephritis.

Few effective therapeutic options are available for treating severe infections caused by extensively drug-resistant Acinetobacter baumannii (XDR-AB). Using a murine thigh-infection model, we examined the in vivo efficacy of colistin in combination with meropenem, tigecycline, fosfomycin, fusidic acid, rifampin, or sulbactam against 12 XDR-AB strains. Colistin, tigecycline, rifampin, and sulbactam monotherapy significantly decreased bacterial counts in murine thigh infections compared with those observed in control mice receiving no treatment. Colistin was the most effective agent tested, displaying bactericidal activity against 91.7% of strains at 48 h post-treatment. With strains showing a relatively low minimum inhibitory concentration (MIC) for meropenem (MIC ≤ 32 mg/L), combination therapy with colistin plus meropenem caused synergistic inhibition at both 24 h and 48 h post-treatment. However, when the meropenem MIC was ≥64 mg/L, meropenem did not significantly alter the efficacy of colistin. The addition of rifampin and fusidic acid significantly improved the efficacy of colistin, showing a synergistic effect in 100% and 58.3% of strains after 24 h of treatment, respectively, while the addition of tigecycline, fosfomycin, or sulbactam did not show obvious synergistic activity. No clear differences in activities were observed between colistin-rifampin and colistin-fusidic acid combination therapy with most strains. Overall, our in vivo study showed that administering colistin in combination with rifampin or fusidic acid is more efficacious in treating XDR-AB infections than other combinations. The colistin-meropenem combination may be another appropriate option if the MIC is ≤32 mg/L. Further clinical studies are urgently needed to confirm the relevance of these findings.

International Normalized Ratio (INR), which standardizes prothrombin time (PT) during oral anticoagulation, has been extended to standardize PT in liver diseases and is included in all prognostic models of survival, the classification of CHILD-Pugh or Meld. However, the mechanisms of PT prolongation in liver diseases differ from those involved in oral anticoagulation. Our aim was to assess the validity of the INR system for patients with liver disease associated with viral hepatitis. We prospectively collected blood samples from 61 patients with liver disease associated with viral hepatitis; control patients were on warfarin (n = 20). PTs were measured on a STA-R coagulometer with six thromboplastin reagents, and INRs were calculated using instrument-specific ISIs. Simultaneously, we selected 15 pairs of patients in the study population and in the control population such that INR values for each patient pair are almost equal. For these 15 pairs of patients, we performed factor assays and measured the coagulant activities of factors II, V, VI, and X and fibrinogen. Analysis of results for the control population confirms the validity of the INR system for patients on oral anticoagulants in that there was no significant difference between the reported INRs for the six different thromboplastin reagents. Conversely, for the study population, there was a significant difference between the INR results using the different reagents. Results for fibrinogen and factors V, VII, and X showed significant differences between the two groups; however, control and patient results for factor II were not statistically different. The INR system is not valid for comparison of patients with liver disease associated with viral hepatitis because different reagents do not yield the same INR for the same sample.

BACKGROUND: Hematology analysis is an essential component of patient assessment and used in screening, diagnosis, and the planning of care. The objective of the study was to find out the suitable hematology review criteria for large scale general hospitals in China via the analysis of experimental data from Sysmex XE-2100 hematology analyzer. METHODS: A total 1486 blood samples were detected with the Sysmes XE-2100. Based on hematology review criteria suggested by international consensus group and a positive smear finding new optimal review rules were determined. RESULTS: With the International Article 41 Review Rules, the true positive ratio (TP), the false positive ratio (FP), the true negative ratio (TN), and the false negative ratio (FN) was 14.0% (208/1486), 31.49% (468/1486) 52.42% (779/1486), and 2.09% (31/1486), respectively. With the help of Laboman 4.2 software (the Sysmex Corporation), 19 rules for review of automated CBC and WBC differential were set up. With our review rules, the TP, FP, TN, and FN was 13.86% (206/1486), 25.17% (374/1486), 58.75% (873/1486) and 2.22% (33/1486), respectively. The review rules were validated, the FN was 0.96%, blasts and immature cells were not omitted. CONCLUSIONS: The review criteria can be developed in light of the rules of the International Consensus Group for Hematology Review, but should be improved depending on different laboratory's requirements.

Several automated urine sediment analyzers have been introduced to clinical laboratories. Automated microscopic pattern recognition is a new technique for urine particle analysis. We evaluated the analytical and diagnostic performance of the UriSed automated microscopic analyzer and compared with manual microscopy for urine sediment analysis.Precision, linearity, carry-over, and method comparison were carried out. A total of 600 urine samples sent for urinalysis were assessed using the UriSed automated microscopic analyzer and manual microscopy.Within-run and between-run precision of the UriSed for red blood cells (RBC) and white blood cells (WBC) were acceptable at all levels (CV20%). Within-run and between-run imprecision of the UriSed testing for cast, squamous epithelial cells (EPI), and bacteria (BAC) were good at middle level and high level (CV20%). The linearity analysis revealed substantial agreement between the measured value and the theoretical value of the UriSed for RBC, WBC, cast, EPI, and BAC (r0.95). There was no carry-over. RBC, WBC, and squamous epithelial cells with sensitivities and specificities were more than 80% in this study.There is substantial agreement between the UriSed automated microscopic analyzer and the manual microscopy methods. The UriSed provides for a rapid turnaround time.

BACKGROUND Microscopic examination is essential for urine analysis, but a time-consuming procedure. This study was undertaken to evaluate an automated urinalysis system - the Sysmex UF-1000i (URISYS 2400) for the analysis of urine constituents including chemistry components and particles. The objective was to screen urine samples and determine the screening criteria which would minimize the number of specimens reviewed with the microscope yet ensuring correct results. METHODS A total of 1300 urine samples were sent for urinalysis using the automated system and compared with results obtained from manual microscopy using the Fuchs-Rosenthal counting chamber. RESULTS Using Pearson statistics, we observed correlation between the UF-1000i and manual microscopy: for red blood cells (RBCs) r was 0.949, for white blood cells (WBCs) r was 0.882, for epithelial cells (EC) r was less than 0.76, for casts r was less than 0.7, while correlation between the URISYS 2400 and manual microscopy: for red blood cells r was 0.772 and for white blood cells r was 0.771. With the help of Uriaccess (an expert system provided by the Sysmex Corporation), 37 rules for microscopic review were set up. The review rules were validated, the review rate was less than 30% and the false-positive and false-negative results were acceptably low. CONCLUSIONS UF-1000i is capable of reproducible measurement of urine particles within the clinically relevant range and shows its advantage over URISYS 2400. It is an optimal strategy for urine sample screening using the combination of the two methods.

To investigate the mechanism of activation of platelet by shear stress.Specimens of whole blood were exposed to shear stress of different intensity (100, 150, 1,000, and 3,000 s(-1)) by modified cone-and-plate rotational viscometer. Then the platelets were stained with fluorescent antibody technique by flow cytometry to examine the levels of four kinds of platelet membrane glycoproteins: PAC-1, CD62P, GPIb/IX, and GPIIb/IIIa.The expression of PAC-1 and CD62P on the surface of unactivated platelets was very low, with the rates of 1.7% +/- 1.1% (n = 5) and 0.9% +/- 0.5% (n = 5) respectively. When the platelets were exposed to high shear stress, the expression of these two kinds of glycoproteins increased. The level of PAC-1 increased quickly and obviously in a short time, reached the peak value, 73.6% +/- 7.4%, in half a minute after exposure to the shear stress of 3,000 s(-1) and decreased soon. The expression rate of CD62P was low when the platelets were exposed to low shear stress, and then increased slowly along with the increase of shear stress. After exposure to the shear stress of 3,000 s(-1) for 7 minutes, the expression rate of CD62P reached 26.4% +/- 3.5%. GPIb/IX and GPIIb/IIIa were found to exist on the surface of more than 96.5% - 99.4% of unactivated platelets. No significant change of GPIb/IX expression was found at relatively low shear stress. When the shear stress increased, the expression rate of GPIb/IX increased to an extent in 1 minute, then decreased continually. The fluorescent intensity of GPIb/IX was 72.4 +/- 6.7 while unactivated, and decreased to 44.9 +/- 4.9 (n = 5) after exposure to the shear stress of 3,000 s(-1) for 7 minutes. The expression of GPIIb/IIIa on surface of unactivated platelets, with fluorescent intensity of 98.5 +/- 12.1, began to increase under low intensity of shear stress. When the stress increased, GPIIb/IIIa expression increased quickly in a short time, and began to decrease in 7 minutes with the fluorescent intensity of 159.5 +/- 23.6 (n = 5) by then.High shear stress causes change of expression of platelet membrane glycoproteins. Platelets are activated by high shear stress directly and independently, and the activation may be mediated by these glycoproteins.

Background HIV trans-activator protein (Tat) is the crucial factor to control HIV transcription, and is usually considered as an important immunogen for the design of HIV vaccine. Recent studies reported some special bio-activities of Tat protein on immunoregulation. However, to date, few studies have focused on exploring the effects of Tat expression plasmid (pTat) on regulating the immune responses induced by HIV DNA vaccines. In this study, our main objective is to investigate the immunoregulation mediated by pTat in mice. Methods Four gene-coding plasmids (pTat, pGag, pEnv and pPol) were constructed, and the gene expression was detected by western blot method. The effects of pTat on regulating the immune responses to antigens Gag, Env, Pol were assessed by enzyme-linked immunospot and enzyme-linked immunosorbent assay. The data was analysed by one-way analysis of variance. Results After two immunizations, mice vaccinated with antigen expressing plasmid (pGag, pEnv or pPol) plus pTat exhibited significantly stronger IFN-gamma response than that vaccinated with the corresponding antigen alone. Moreover, mice receiving two injections of antigen plus pTat exhibited the same strong IFN-gamma response as those receiving three injections of antigen alone did. Furthermore, addition of pTat not only induced a more balanced Th1 and Th2 response, but also broadened IgG subclass responses to antigens Gag and Pol. Conclusion pTat exhibited the appreciable effects on modulating immune responses to HIV antigens Gag, Env and Pol, providing us interesting clues on how to optimize HIV DNA vaccine.