Accurate Clinical Diagnosis of Liver Cancer Based on Simultaneous Detection of Ternary Specific Antigens by Magnetic Induced Mixing Surface-Enhanced Raman Scattering Emissions.

Establishing an accurate, simple, and rapid serodiagnosis method aiming for specific cancer antigens is critically important for the clinical diagnosis, therapy, and prognostication of cancer. Currently, surface-enhanced Raman scattering (SERS) readout techniques challenge fluorescent-based detection methods in terms of both optical stability and more importantly multiple detection capability, which become more desirable for clinical diagnostics. We thus started using an interference-free mixing SERS emission (m-SERS) readout to simultaneously indicate, for the first time, three specific liver cancer antigens, including α-fetoprotein (AFP), carcinoembryonic antigen (CEA), and ferritin (FER), even in one clinical serum sample. Here, three triple bonds (C≡N and C≡C) coded SERS tags contribute separate SERS emissions located at 2105, 2159, and 2227 cm-1, respectively; must have one-to-one correspondence from AFP, to FER, to CEA, In the process of detection, the mature double antibody sandwich allows the formation of microscale core-satellite assembly structure between a magnetic bead (MB) and single SERS tags, and therefore a pure and single SERS emission can be observed under the routine excitation laser spot. Because of the action of magnetic force, the uniform 3D packing of SERS tags absorbed MBs will in contrast generate a so-called m-SERS signals. With the help of enrichment and separation by MBs, the proposed m-SERS immunoassay provides an extremely rapid, sensitive, and accurate solution for multiplex detection of antigens or other biomarkers. Herein, the limit of detection (LOD) for simultaneous m-SERS detection of AFP, CEA, and FER was 0.15, 20, and 4 pg/mL, respectively. As expected for 39 clinical serum samples, simultaneous detection of ternary specific antigens can significantly improve the accuracy of liver cancer diagnosis.

Splicing Nanoparticles-Based "Click" SERS Could Aid Multiplex Liquid Biopsy and Accurate Cellular Imaging.

Here, a completely new readout technique, so-called "Click" SERS, has been developed based on Raman scattered light splice derived from nanoparticle (NP) assemblies. The single and narrow (1-2 nm) emission originating from triple bond-containing reporters undergoes dynamic combinatorial output, by means of controllable splice of SERS-active NPs analogous to small molecule units in click chemistry. Entirely different to conventional "sole code related to sole target" readout protocol, the intuitional, predictable and uniquely identifiable "Click" SERS is relies on the number rather than the intensity of combinatorial emissions. By this technique, 10-plex synchronous biomarkers detection under a single scan, and accurate cellular imaging under double exposure have been achieved. "Click" SERS demonstrated multiple single band Raman scattering could be an authentic optical analysis method in biomedicine.

Alkyne-Modulated Surface-Enhanced Raman Scattering-Palette for Optical Interference-Free and Multiplex Cellular Imaging.

The alkyne tags possess unique interference-free Raman emissions but are still hindered for further application in the field of biochemical labels due to its extremely weak spontaneous Raman scattering. With the aid of computational chemistry, herein, an alkyne-modulated surface-enhanced Raman scattering (SERS) palette is constructed based on rationally designed 4-ethynylbenzenethiol derivatives for spectroscopic signature, Au@Ag core for optical enhancement and an encapsulating polyallylamine shell for protection and conjugation. Even for the pigment rich plant cell (e.g., pollen), the alkyne-coded SERS tag can be highly discerned on two-dimension distribution impervious to strong organic interferences originating from resonance-enhanced Raman scattering or autofluorescence. In addition, the alkynyl-containing Raman reporters contribute especially narrow emission, band shift-tunable (2100-2300 cm(-1)) and tremendously enhanced Raman signals when the alkynyl group locates at para position of mercaptobenzene ring. Depending on only single Raman band, the suggested alkyne-modulated SERS-palette potentially provides a more effective solution for multiplex cellular imaging with vibrant colors, when the hyperspectral and fairly intense optical noises originating from lower wavenumber region (<1800 cm(-1)) are inevitable under complex ambient conditions.

The small silver nanoparticle-assisted homogeneous sensing of thiocyanate ions with an ultra-wide window based on surface-enhanced Raman-extinction spectroscopy.

For the first time, we present an original sensing strategy with an ultra-wide detection window from 17 nM to 20 mM to detect SCN- ions. Initially, we investigated the clustering and optical properties of noble metal sol nanoparticles (NPs) due to the competitive interaction of thiocyanate ions (SCN-) and cetyltrimethylammonium bromide (CTAB) under weak acidic conditions, and found that different dimensions and scales of nanoclusters containing the alkyne-embedded Au@Ag NPs and relatively small Ag NPs could be achieved by the mediation of CTAB through electrostatic forces and hydrophobic interaction, in which SCN- could be covalently bonded with the silver surface of NPs to form a compact molecular layer (-Ag-S-C[triple bond, length as m-dash]N), and CTAB could only occupy remaining sites. In this process, we found that SCN- always runs counter to CTAB and tends to dissolve nanoclusters, so that they occupy the exposed surface of NPs in nanoclusters rather than the binding sites of one another. Remarkably, when the concentration of SCN- initially increased, two highly recognizable SERS emissions, which were assigned to alkyne reporter molecules (2208 cm-1) and C[triple bond, length as m-dash]N of SCN- (2110 cm-1), respectively, were rapidly detected, and their ratios (I2110/I2208) increased linearly proportional to the concentration of SCN- over a range of 17 nM to 172 µM, with a limit of detection (LOD) of 10 nM. With the further increase of SCN-, small Ag NPs started to desorb from the surface of individual Au@Ag NPs and dissociated in the solution but did not contribute to SERS signals. Instead, the surface plasmon resonance (SPR) peak of pure silver NPs at 385 nm increased gradually in the range from 0.5 to 20 mM with an LOD of 0.2 mM. Of particular significance, this simple sensor in conjunction with surface-enhanced Raman-extinction spectroscopy can be used for the rapid detection of extensive samples with an ultra-wide detection window, such as body fluids (saliva, urine, and serum) and food (milk powder and brassica vegetables), which is far superior to that of ion chromatography (IC).

Rapid and Reliable Detection of Alkaline Phosphatase by a Hot Spots Amplification Strategy Based on Well-Controlled Assembly on Single Nanoparticle.

The first appeal of clinical assay is always accurate and rapid. For alkaline phosphatase (ALP) monitoring in medical treatment, a rapid, reliable surface-enhanced Raman scattering (SERS) test kit is designed based on a "hot spots" amplification strategy. Consisting of alkyne-tagged Au nanoparticles (NPs), Ag+, and enzyme substrate, the packaged test kit can achieve one-step clinical assay of ALP in human serum within several minutes, while the operation is simple as it directly inputs the sample into the test kit. Here, Ag+ ions are adsorbed onto the surface of Au core due to electrostatic interaction between Ag+ and the negatively charged donor surface, then enzymatic biocatalysis of ALP triggers the reduction of Ag+ and subsequently silver growth occurs on every Au core surface in a controllable manner, forming "hot spots" between the Au core and Ag shell, in which the SERS signal of alkyne Raman reporters would be highly amplified. Meanwhile, ALP mediates a redox reaction of Ag+ as well as the dynamic silver coating process so the increase of SERS intensity is well-controlled and can be recognized with increasing amounts of the targets. Instead of conventional NP aggregation, this leads to a more reproducible result. In particular, the distinct Raman emission from our self-synthesized alkyne reporter is narrow and stable with zero background in the Raman silent region, suffering no optical fluctuation from biosystem inputs and the detection results are therefore reliable with a limit of detection of 0.01 U/L (2.3 pg/mL). Along with ultrahigh stability, this SERS test kit therefore is an important point-of-care candidate for a reliable, efficacious, and highly sensitive detection method for ALP, which potentially decreases the need for time-consuming clinical trials.

Chromosomal imbalances in nasopharyngeal carcinoma: a meta-analysis of comparative genomic hybridization results.

Nasopharyngeal carcinoma (NPC) is a highly prevalent disease in Southeast Asia and its prevalence is clearly affected by genetic background. Various theories have been suggested for its high incidence in this geographical region but to these days no conclusive explanation has been identified. Chromosomal imbalances identifiable through comparative genomic hybridization may shed some light on common genetic alterations that may be of relevance to the onset and progression of NPC. Review of the literature, however, reveals contradictory results among reported findings possibly related to factors associated with patient selection, stage of disease, differences in methodological details etc. To increase the power of the analysis and attempt to identify commonalities among the reported findings, we performed a meta-analysis of results described in NPC tissues based on chromosomal comparative genomic hybridization (CGH). This meta-analysis revealed consistent patters in chromosomal abnormalities that appeared to cluster in specific "hot spots" along the genome following a stage-dependent progression.

Downregulation of wild-type p53 protein by HER-2/neu mediated PI3K pathway activation in human breast cancer cells: its effect on cell proliferation and implication for therapy.

Overexpression and activation of HER-2/neu (also known as c-erbB-2), a proto-oncogene, was found in about 30% of human breast cancers, promoting cancer growth and making cancer cells resistant to chemo- and radio-therapy. Wild-type p53 is crucial in regulating cell growth and apoptosis and is found to be mutated or deleted in 60-70% of human cancers. And some cancers with a wild-type p53 do not have normal p53 function, suggesting that it is implicated in a complex process regulated by many factors. In the present study, we showed that the overexpression of HER-2/neu could decrease the amount of wild-type p53 protein via activating PI3K pathway, as well as inducing MDM2 nuclear translocation in MCF7 human breast cancer cells. Blockage of PI3K pathway with its specific inhibitor LY294002 caused G1-S phase arrest, decreased cell growth rate and increased chemo- and radio-therapeutic sensitivity in MCF7 cells expressing wild-type p53. However, it did not increase the sensitivity to adriamycin in MDA-MB-453 breast cancer cells containing mutant p53. Our study indicates that blocking PI3K pathway activation mediated by HER-2/neu overexpression may be useful in the treatment of breast tumors with HER-2/neu overexpression and wild-type p53.

[The effect of HER2/neu overexpression on p53 gene expression, cell proliferation and sensitivity to gamma-irradiation via the PI3K/Akt pathway in breast cancer cell MCF7].

OBJECTIVE: To investigate the effect of HER2/neu overexpression on the wild p53 gene expression, cell proliferation and sensitivity to gamma-irradiation via phosphatidylinositol 3-kinase (PI3K) pathway in human breast cancer cell MCF7. METHODS: Lipofectin-mediated gene transfection method was used to transfer HER2/neu into MCF7 cells. Expression of HER2/neu, p53, Akt and p-Akt protein after PI3K pathway inhibitor LY294002 treatment was determined by Western blot. Cell proliferation and cell surviving fraction after gamma-irradiation treatment were assayed by MTT. RESULTS: Eighteen of HER2/neu stably transfected MCF7 cell clones were established, one of them was HER2/neu overexpressing. HER2/neu overexpressing MCF7 cells showed higher p-Akt expression and lower p53 expression than those of parental MCF7 cells, which could be abrogated by LY294002. HER2/neu overexpressing MCF7 cells had higher proliferation rate and lower sensitivity to gamma-irradiation than those of parental MCF7 cells, which could be opposed by LY294002. CONCLUSION: Overexpression of HER2/neu induces reduced expression of wild-type p53 protein, relatively high cell proliferation and low sensitivity to gamma-irradiation in breast cancer cell MCF7 by activating PI3K/Akt pathway, which may contribute to therapeutic resistance in some breast cancer patients with wild-type p53 gene status.

[Correlation study of expression levels of prostate-specific membrane antigen and prostate-specific antigen with Gleason score of prostate carcinoma].

OBJECTIVE: To study the correlation of prostate-specific membrane antigen (PSMA) and prostate-specific antigen (PSA) expression with Gleason score of prostate carcinoma. METHODS: Monoclonal antibodies against epitopes of PSMA extracellular domain were prepared, with which the expression of PSMA of prostate carcinoma (PC) was determined by immunohistochemical staining. Correlation of its expression with Gleason score of PC was statistically analyzed, and compared with that of PSA. RESULTS: Eight hybridoma cell lines secreting monoclonal antibodies specific for PSMA were prepared. PSMA expression level was positively correlated with Gleason score. In poorly differentiated prostate carcinoma, the expression intensity of PSMA was higher than that of medium-and well-differentiated prostate carcinoma (P < 0.01). However, there was no correlation between level of PSA expression and Gleason score (P > 0.05). CONCLUSION: PSMA expression level may be used as a useful surrogate marker in Gleason grading of prostate carcinoma. It may be a more suitable target than PSA in antibody mediated immunotherapy against poorly differentiated prostate carcinoma which is usually not sensitive to hormonal therapy.

[Overexpression of HER2/neu downregulates wild p53 protein expression via PI3K and Ras/Raf/MEK/ERK pathways in human breast cancer cells].

OBJECTIVE: To evaluate the effect of HER2/neu overexpression on wild p53 protein expression and to delineate the related signal pathways. METHODS: Lipofectin method was used to transfer HER2/neu into human breast tumor cell line MCF7. Overexpression of HER2/neu was then determined by Western blot. Western blot was also used to detect the quantity of p53, Akt, p-Akt, p-Raf, p-MEK, p-ERK protein. Semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) was employed to detect p53 mRNA expression. PI3K pathway inhibitor LY294002 and MEK inhibitor U0126 were used to block the two pathways. The subsequent effect on p53 protein expression was then determined. RESULTS: HER2/neu-overexpressed MCF7 clone (MCF7-neu3) was successfully established, in which the amount of HER2/neu protein was 13 times more than that in parental MCF7 cells. The amount of p53 protein in MCF7-neu3 was 40% of that in parental MCF7 cells (P < 0.01), while there was no difference on p53 mRNA level. There were 2.5, 2.0, 1.6 and 1.6 fold increase in the amount of p-Akt, p-Raf, p-MEK, p-ERK protein respectively in MCF7-neu3 to that in parental MCF7 cells (P < 0.01). When treated with LY294002 or U0126 for 24 hours, the amount of wild p53 protein in MCF7-neu3 cells was 1.7 or 1.5 times higher than those in DMSO treated cells. There were 4.7 or 5.3 times increase in the p53 protein when MCF7-neu3 cells were treated with LY294002 or U0126 for 48 hours (P < 0.01). Similar results were not seen in MDA-MB-453 cells which contained mutant p53. CONCLUSIONS: HER2/neu overexpression can activate PI3K and Ras/Raf/MEK/ERK pathways, resulting in reduction of wild p53 protein expression. This may be the molecular mechanism responsible for the poor prognosis and therapeutic non-responsiveness in HER2/neu-overexpressed breast cancer patients.