Determination of L-selectin in blood plasma using DNA aptamer-based surface-enhanced Raman spectroscopy assay.

In the human body, tumor cell occurrence can be indirectly monitored using the L-selectin concentration in the blood, since selectin ligands are present on the surface of tumor cells, and with tumor progression, a decrease in L-selectin levels can be expected and observed. In this study, we present a selective DNA-based surface-enhanced Raman spectroscopy (SERS) assay for the detection and determination of L-selectin in biological samples. Two calibration curves (linear in the 40-190 ng mL-1 region and exponential in the 40-500 ng mL-1 region) are fitted to the obtained SERS experimental data, i.e., the ratio of I732/I1334 band intensities (LOQ = 46 ng mL-1). Calculated determination coefficients are found to be R2 = 0.997 for the linear region of the calibration curve and R2 = 0.977 for the exponential region. Moreover, we demonstrate very good selectivity of the assay even in the presence of P- and E-selectin in a sample containing L-selectin. With our SERS assay, the L-selectin concentration in biological samples can be estimated directly from the calibration curves.

Lung Cancer: Spectral and Numerical Differentiation among Benign and Malignant Pleural Effusions Based on the Surface-Enhanced Raman Spectroscopy.

We present here that the surface-enhanced Raman spectroscopy (SERS) technique in conjunction with the partial least squares analysis is as a potential tool for the differentiation of pleural effusion in the course of the cancerous disease and a tool for faster diagnosis of lung cancer. Pleural effusion occurs mainly in cancer patients due to the spread of the tumor, usually caused by lung cancer. Furthermore, it can also be initiated by non-neoplastic diseases, such as chronic inflammatory infection (the most common reason for histopathological examination of the exudate). The correlation between pleural effusion induced by tumor and non-cancerous diseases were found using surface-enhanced Raman spectroscopy combined with principal component regression (PCR) and partial least squares (PLS) multivariate analysis method. The PCR predicts 96% variance for the division of neoplastic and non-neoplastic samples in 13 principal components while PLS 95% in only 10 factors. Similarly, when analyzing the SERS data to differentiate the type of tumor (squamous cell vs. adenocarcinoma), PLS gives more satisfactory results. This is evidenced by the calculated values of the root mean square errors of calibration and prediction but also the coefficients of calibration determination and prediction (R2C = 0.9570 and R2C = 0.7968), which are more robust and rugged compared to those calculated for PCR. In addition, the relationship between cancerous and non-cancerous samples in the dependence on the gender of the studied patients is presented.

SERS-based sensor for direct L-selectin level determination in plasma samples as alternative method of tumor detection.

Selectin ligands are present on the surface of tumor cells, for this reason lowering the L-selectin level in the blood and lymph can indicate presence of the tumor. Therefore the selectin level in the plasma are potential targets for anticancer therapy. We demonstrate the surface enhanced Raman spectroscopy (SERS)-based sensor for the determination of L-selectin level in biological samples that can be used in medical diagnosis. The combination of SERS with the method of multivariate analysis as principle component analysis (PCA) allows to strengthen the presented data analysis. The loadings of PCA permit to indicate those vibration modes, that are the most important for the assumed identification (bands at 1574, 1450, 1292 cm-1 ). Two bands at 1286 and 1580 cm-1 were selected for the determination of the calibration curve (bands intensities I1286 /I1580 ratio). The L-selectin level of biological samples can be read, directly from the calibration curve. The presented sensor is as a sensitive tool with good specificity and selectivity of L-selectin, even in the case of coexistence of P- and E-selectin.

Brain tumour homogenates analysed by surface-enhanced Raman spectroscopy: Discrimination among healthy and cancer cells.

One of the biggest challenge for modern medicine is to make a discrimination among healthy and cancerous tissues. Therefore, nowadays big effort of scientist are devoted to find a new way for as fast as possible diagnosis with as much as possible accuracy in distinguishing healthy from cancerous tissues. That issues are probably the most important in the case of brain tumours, when the diagnosis time plays a great role. Herein we present the surface-enhanced Raman spectroscopy (SERS) together with the principal component analysis (PCA) used to identify the spectra of different brain specimens, healthy and tumour tissues homogenates. The presented analyses include three sets of brain tissues as control samples taken from healthy objects (one set consists of samples from four brain lobes and both hemispheres; eight samples) and the brain tumours from five patients (two Anaplastic Astrocytoma and three Glioblastoma samples). Results prove that tumour brain samples can be discriminated well from the healthy tissues by using only three main principal components, with 96% of accuracy. The largest influence onto the calculated separation is attributed to the spectral regions corresponding in SERS spectra to vibrations of the L-Tryptophan (1450, 1278 cm-1), protein (1300 cm-1), phenylalanine and Amide-I (1005, 1654 cm-1). Therefore, the presented method may open the way for the probable application as a very fast diagnosis tool alternative for conventionally used histopathology or even more as an intraoperative diagnostic tool during brain tumour surgery.

Comparative study of molecular recognition of folic acid subunits with cyclodextrins.

The complexation of pteroic acid and pterine, subunits of folic acid, with native cyclodextrins (α‒, ß‒, and γ‒CDs) was studied in solution (UV-vis), and in the solid state (thermal analysis, IR and Raman). UV-vis titrations at pH = 7.4 provided data regarding stoichiometry of the formed complexes as well as their associations constants. Stability of the complexes increases in the series: γ‒CD < ß‒CD << α‒CD for pterine, and γ‒CD < α‒CD << ß‒CD for pteroic acid. The mode of complexation was further exploited by molecular modeling studies (docking studies, PM6) showing additionally changes in conformation of pteroic acid upon complexation. Comparison of the association constants of the complexes of pterine and pteroic acid with native cyclodextrins with data obtained for analogous complexes with folic acid shows that all folic acid complexes are less stable than those formed from its subunits.

Structural diversity in the host-guest complexes of the antifolate pemetrexed with native cyclodextrins: gas phase, solution and solid state studies.

The complexation of the antifolate pemetrexed (PTX) with native cyclodextrins was studied. This process, along with the findings gathered for the structurally related folic acid was treated as a model for exploiting host-guest interactions of this class of guest molecules in the gas phase, in solution and in the solid state. Mass spectrometry was employed for the investigation of the architecture and relative gas-phase stabilities of these supramolecular complexes. The mode of complexation was further tracked by 1D and 2D NMR proving the formation of the exclusion-type complex with α-CD and pseudorotaxane inclusion-type complexes with ß-, and γ-CDs. UV-vis titrations at pH 7.4 gave association constants for the obtained complexes. The stability of the complexes increases in the series: α-CD/PTX < γ-CD/PTX << ß-CD/PTX. The association of PTX with a monomer cyclodextrin equivalent - methyl α-D-glucopyranoside - was investigated for a deeper understanding of the type of host-guest interactions. Solid state studies of PTX/CDs were performed using FTIR-ATR and Raman spectroscopy techniques.

ZnO oxide films for ultrasensitive, rapid, and label-free detection of neopterin by surface-enhanced Raman spectroscopy.

Efficient and low-cost surface-enhanced Raman scattering (SERS) substrates based on Au coated zinc oxide layers for the detection of neopterin were prepared. These substrates showed high sensitivity to p-mercaptobenzoic acid (p-MBA) at a low concentration of 10(-9) M and an enhancement factor of over 10(7) was achieved. The uniform density of SERS-active "hot-spots" on a Si/ZnO/Au surface results in high reproducibility towards detecting p-MBA at 50 different, randomly selected positions on a single substrate (RSD = 9%) and on six different SERS substrates prepared under identical conditions (RSD = 11%). These SERS substrates show good performance in the detection of neopterin, a biologically important molecule whose concentration levels reflect the stage of activation of the cellular immune system, which is of value in the studies of pathogenesis and progression of various diseases. The detection limit is found to be as low as 1.4 nmol L(-1) in blood plasma, which is comparable to that of classic ELISA methods. The average relative standard deviation (RSD) of the proposed method is less than 10%. Moreover, this label-free strategy of detection gives exact results over a large range, reflecting clinically relevant neopterin concentrations in body fluids. The detection and quantification of neopterin levels in blood or urine might be useful in clinical practice for monitoring the disease activity during treatment and for early detection of many infections and autoimmune, inflammatory, and malignant diseases.