Yingchang Zou

CEA, SCC and NSE levels in exhaled breath condensate—possible markers for early detection of lung cancer

Lung cancer (LC) is the leading cause of cancer-related death. The sensitive and non-invasive diagnostic tools in the early stage are still poor. We present a pilot study on the early diagnosis of LC by detecting markers in exhaled breath condensate (EBC). EBC samples were collected from 105 patients with LC and 56 healthy controls. We applied chemiluminescence immunoassay to detect CEA (carcinoembryonic antigen), SCC (squamous cell carcinoma) antigen and NSE (neuron specific enolase) in EBC and serum. Concentrations of markers were compared between independent groups and subgroups. A significantly higher concentration level of each marker was found in patients with LC than healthy controls. The areas under curve of receiver operating characteristic (ROC) curves were 0.800, 0.771, 0.659, 0.679, 0.636 and 0.626 for EBC-CEA, serum-CEA, EBC-SCC, serum-SCC, EBC-NSE and serum-NSE, respectively. Markers in EBC had a higher positive rate (PR) and were more specific to histologic types than markers in serum. In addition, multivariate analysis was performed to evaluate the association of presenting markers with the stages of non-small cell lung cancer (NSCLC). EBC-CEA showed the best predictive characteristic (p < 0.006) of early-NSCLC. Our study suggested that tumor markers in EBC may have a better diagnostic performance for LC than those in serum. With further investigation on the combination of markers in EBC, detection of EBC could probably be a novel and non-invasive method to detect NSCLC earlier.

Optimization of volatile markers of lung cancer to exclude interferences of non-malignant disease.

BACKGROUND Breath analysis became promising for noninvasive diagnosis of cancer with sophisticated spectrometry technology introduced. OBJECTIVE This study aimed to select volatile markers for lung cancer detection, which exclude the influences from non-malignant lung diseases. METHODS 171 subjects who were divided into three groups: patients with LC, patients with PNMD and healthy controls were enrolled in our studies as training cohort. The volatile organic compounds (VOCs) in their breath samples were analyzed with solid-phase micro-extraction/gas chromatography/mass spectrometry (SPME-GCMS). Markers were selected by receiver operating characteristic (ROC) curves. After that, 78 subjects with high morbidity of LC were employed as validation cohort. Their breath samples were analyzed by thermal desorption instrument/gas chromatography/mass spectrometry (TD-GCMS). RESULTS Through a series of comparisons among lung cancer patients, pulmonary non-malignant diseases patients, and healthy participants in training cohort, Nonane,5-(2-methyl-)propyl-; phenol,2,6-di-tert-butyl-,4-methyl-; dodecane,2,6,11-trimethyl-; hexadecanal and pentadecane,8-hexyl- were selected as markers for lung cancer diagnosis. Principal component analysis was employ to process data from validation cohort. As results, satisfied distinctions have been obtained with detection of these five selected markers, although the detection method is not identical with that used for training cohort. CONCLUSIONS In conclusion, with optimization method described in this paper, breath test could be an effective method for diagnosis of lung cancer and avoid the interference of pulmonary non-malignant diseases.

A novel sensitive cell-based Love Wave biosensor for marine toxin detection

A novel HepG2 cell-based biosensor using Love Wave sensor was developed to implement the real-time and sensitive detection of a diarrheic shellfish poisoning (DSP) toxin, Okadaic acid (OA). Detachable Love Wave sensor unit and miniaturized 8-channel recording instrument were designed for the convenient experimental preparation and sensor response signal measurement. The Love Wave sensor, whose synchronous frequency is around 160 MHz, was fabricated with ST-cut quartz substrate. To establish a cell-based biosensor, HepG2 cells as sensing elements were cultured onto the Love Wave sensor surface, and the cell attachment process was recorded by this biosensor. Results showed this sensor could monitor the cell attachment process in real time and response signals were related to the initial cell seeding densities. Furthermore, cell-based Love Wave sensor was treated with OA toxin. This biosensor presented a good performance to various OA concentrations, with a wide linear detection range (10-100 μg/L). Based on the ultrasensitive acoustic wave platform, this cell-based biosensor will be a promising tool for real-time and convenient OA screening.

Breath analysis for noninvasively differentiating Acinetobacter baumannii ventilator-associated pneumonia from its respiratory tract colonization of ventilated patients.

A number of multiresistant pathogens including Acinetobacter baumannii (A. baumannii) place a heavy burden on ventilator-associated pneumonia (VAP) patients in intensive care units (ICU). It is critically important to differentiate between bacterial infection and colonization to avoid prescribing unnecessary antibiotics. Quantitative culture of lower respiratory tract (LRT) specimens, however, requires invasive procedures. Nowadays, volatile organic compounds (VOCs) have been studied in vitro and in vivo to identify pathogen-derived biomarkers. Therefore, an exploratory pilot study was conceived for a proof of concept that the appearance and level of A. baumannii-derived metabolites might be correlated with the presence of the pathogen and its ecological niche (i.e. the infection and colonization states) in ICU ventilated patients. Twenty patients with A. baumannii VAP (infection group), 20 ventilated patients with LRT A. baumannii colonization (colonization group) and 20 ventilated patients with neurological disorders, but without pneumonia or A. baumannii colonization (control group) were enrolled in the in vivo pilot study. A clinical isolate of A. baumannii strains was used for the in vitro culture experiment. The adsorptive preconcentration (solid-phase microextraction fiber and Tenax(®) TA) and analysis technique of gas chromatography-mass spectrometry were applied in the studies. Breath profiles could be visually differentiated between A. baumannii cultivation in vitro and culture medium, and among in vivo groups. In the in vitro experiment, nine compounds of interest (2,5-dimethyl-pyrazine, 1-undecene, isopentyl 3-methylbutanoate, decanal, 1,3-naphthalenediol, longifolene, tetradecane, iminodibenzyl and 3-methyl-indene) in the headspace were found to be possible A. baumannii derivations. While there were eight target VOCs (1-undecene, nonanal, decanal, 2,6,10-trimethyl-dodecane, 5-methyl-5-propyl-nonane, longifolene, tetradecane and 2-butyl-1-octanol) exhibiting characteristics of A. baumannii VAP derivations. The selected VOC profile in vivo could be adopted to efficiently differentiate the presence of LRT A. baumannii from its absence, and LRT A. baumannii infection from its colonization (AUC  =  0.89 and 0.88, respectively). It is not feasible to simply transfer the metabolic biomarkers from the in vitro condition to in vivo. The direct detection of exhaled A. baumannii-derived VOCs may be adopted for an early alert of the LRT bacterial presence in ventilated ICU patients, and even in different parasitic states of A. baumannii (i.e. infection and colonization). However, further refinement and validation are required before its clinical use.

A point-of-care testing system with Love-wave sensor and immunogold staining enhancement for early detection of lung cancer

It has been reported that detection of exhaled breath condensate (EBC) is available for studies of pulmonary diseases, especially lung disease. In order to detect lung cancer (LC) at early stage, a point-of-care testing system suitable for measurement of tumor markers in EBC is developed. The assay, based on gold nanoparticle sandwich immunoassay and subsequent gold staining, was performed on a Love-wave sensor packaged inside a chip cartridge. Benefit from high sensitivity of Love-wave sensor, oriented immobilization of coating antibodies and immunogold staining enhancement, the present immunosensor could provide a sensitive, specific and rapid measurement. Carcinoembryonic antigen (CEA), neuron specific enolase (NSE) and squamous cell carcinoma antigen (SCC) in EBC collected from 17 patients with LC and 13 healthy volunteers were detected by this system. Results were compared with commercial chemiluminescence immunoassay and showed high correlation between two methods. Additionally, it revealed significantly statistical differences existing between two groups of subjects. These results indicate that the present system is suitable for detection of tumor markers in EBC and could be used as assistant tools for early detection of LC.

Electronic Nose and Electronic Tongue

The sensations of smell and taste resulting from a series of specific and nonspecific molecular recognition can be used as an analytical tool in many industries to measure the quality of food, drinks, and chemical products. In a few cases, there are olfactory receptor s or gustatory receptors which are specific for individual chemical molecules. However, most tastes and odorants are identified through a synthesis of the global chemical information from nonspecific interactions. Taking mammalian gustation as an example, the combination of “gustatory buds” which respond to five taste categories : sour , sweet , bitter , salty , and umami creates a distinct pattern for each taste.

A novel Love Wave biosensor for rapid and sensitive detection of marine toxins

Marine toxins are produced by plankton and do a great harm to human through food chain by accumulating in shellfishes and fishes. It is highly required and favorable to develop novel methods for the rapid and efficient detection of marine toxins to avoid the poisoning cases that have occurred frequently in many countries. This study presents a real-time Love Wave biosensor for the rapid detection of okadaic acid (OA), which used HepG2 cell lines as the sensing elements. The results indicate that this cell-based biosensor can provide real-time information of cellular activities induced by okadaic acid and has a higher sensitivity than the conventional cell-based assay. It is suggested that this cell-based biosensor can be used as a convenient and efficient method for marine toxin detection, which has a great potential to contribute to avoid the harmful effects of marine toxins on the human health.

Confounding effect of benign pulmonary diseases in selecting volatile organic compounds as markers of lung cancer

Lung cancer (LC) is a leading cause of cancer-related morbidity and mortality globally, and exhaled breath testing has been considered as a fast, convenient and non-invasive way to diagnose LC in its early stages. Volatile organic compounds (VOCs), as markers of LC in exhaled breath, have been widely investigated for cancer diagnosis. However, few studies have reported on the interference of benign pulmonary diseases (BPD) in the selection of VOC markers for LC. During this study, 207 samples were analyzed using thermal desorption instrumentation/gas chromatography/mass spectrometry (TD-GCMS) to detect C6-C30 VOCs, and all samples were divided into four groups: LC group, BPD group, lung disease (LD) group (including LC group and BPD group) and healthy group. To make up for the deficiency of detecting low carbon hydrocarbons (<C6), 277 samples were analyzed using solid-phase micro-extraction/gas chromatography/mass spectrometry (SPME-GCMS), divided among the four groups. VOC markers were selected by reference to the receiver operating characteristics curve. With the comparisons among the LC group, BPD group and healthy group from TD-GCMS and SPME-GCMS results, we found that exhaled VOCs are capable of discriminating LC group versus healthy group and BPD group versus healthy group with a consistency of 70%-80%. However, no VOCs can be selected with good discrimination capability between the LC group and BPD group, indicating that BPD interferes significantly in VOC marker selection for LC. To discriminate breath samples from the LD group and healthy group, 11 VOCs, including ten selected from TD-GCMS and one from SPME-GCMS, were chosen as markers for LD diagnosis. The sensitivity, specificity and overall accuracy of the diagnostic model established using ten VOCs were 80.8%, 84% and 82.7%, and those of the model established by using one VOC were 75.6%, 78.9% and 76.7%. These results validate that LD patients can be effectively discriminated and diagnosed using exhaled VOC analysis. (Clinical trial registration number: ChiCTR-DCD-15007106.).

Bionic 3D spheroids biosensor chips for high-throughput and dynamic drug screening

To perform the drug screening, planar cultured cell models are commonly applied to test efficacy and toxicity of drugs. However, planar cultured cells are different from the human 3D organs or tissues in vivo. To simulate the human 3D organs or tissues, 3D spheroids are developed by culturing a small aggregate of cells which reside around the extracellular matrix and interact with other cells in liquid media. Here we apply lung carcinoma cell lines to engineer the 3D lung cancer spheroid-based biosensor using the interdigitated electrodes for drug efficacy evaluation. The results show 3D spheroid had higher drug resistance than the planar cell model. The anticarcinogen inhibition on different 3D lung cancer spheroid models (A549, H1299, H460) can be quantitatively evaluated by electric impedance sensing. Besides, we delivered combination of anticarcinogens treatments to A549 spheroids which is commonly used in clinic treatment, and found the synergistic effect of cisplatin plus etoposide had higher drug response. To simultaneously test the drug efficacy and side effects on multi-organ model with circulatory system, a connected multiwell interdigitated electrode arraywas applied to culture different organoid spheroids. Overall, the organization of 3D cancer spheroids-based biosensor, which has higher predictive value for drug discovery and personalized medicine screening, is expected to be well applied in the area of pharmacy and clinical medicine.