Label-free electrochemical immunosensor based on graphene/methylene blue nanocomposite

For the specificity of prostate cancer markers, prostate specific antigen (PSA) has been widely used in prostate cancer screening, diagnosis, and treatment after monitoring. In normal male serum, PSA can only be detected in traces of 0-4 ng mL(-1). In this paper, we constructed an electrochemical immunosensor for PSA detection using a nanocomposite film of graphene sheets-methylene blue-chitosan (GS-MB-CS) as electrode material. The nanocomposite film showed high binding affinity to the electrode and was used to immobilize the antibody of PSA. The modification procedure was monitored by cyclic voltammetry (CV). An amperometric biosensor was easily developed based on the response of peak current to the capture of PSA induced by specific antigen-antibody reactions. Under optimum conditions, the amperometric signal decreased linearly with PSA concentration (0.05-5.00 ng mL(-1)). A low limit of detection (13 pg mL(-1)) and a high selectivity are obtained. Moreover, the prepared immunosensor was applied for the analysis of PSA in serum samples with satisfactory results. The proposed method may have a promising future in biochemical assays for high selectivity, good reproducibility, and stability.     

Facile fabrication of magnetic gold electrode for magnetic beads-based electrochemical immunoassay: application to the diagnosis of Japanese encephalitis virus

A novel magnetic beads-based electrochemical immunoassay strategy has been developed for the detection of Japanese encephalitis virus (JEV). The magnetic gold electrode was fabricated to manipulate magnetic beads for the direct sensing applications. Gold-coated magnetic beads were employed as the platforms for the immobilization and immunoreaction process, and horseradish peroxidase was chosen as an enzymatic tracer. The proteins (e.g., antibodies or immunocomplexes) attached on the surface of magnetic beads were found to induce a significant decline in their electric conductivity. Multiwalled carbon nanotubes were introduced to improve sensitivity of the assay. The envelope (E) protein, a major immunogenic protein of JEV, was utilized to optimize the assay parameters. Under the optimal conditions, the linear response range of E protein was 0.84 to 11,200 ng/mL with a detection limit of 0.56 ng/mL. When applied for detection of JEV, the proposed method generated a linear response range between 2×10(3) and 5×10(5) PFU/mL. The detection limit for JEV was 2.0×10(3) PFU/mL, which was 2 orders of magnitude lower than that of immunochromatographic strip and similar to that obtained from RT-PCR. This method was also successfully applied to detect JEV in clinical specimens.     

Facile and rapid magnetic relaxation switch immunosensor for endocrine-disrupting chemicals

In order to develop facile, fast and sensitive detection methods for endocrine-disrupting chemicals (EDCs), we described a sensitive biosensing system involving magnetic relaxation switch, based on the assembly of cross-linked superparamagnetic iron oxide (CLIO) nanoparticles induced by the antigen-antibody biorecognition. The design of smart CLIO-based superparamagnetic iron oxide nanoparticles and antigen-OVA was described for the detection of bisphenol A [2,2-bis (4-hydroxyphenol) propane (BPA)]. The addition of BPA to the rapid magnetic relaxation switch immunosensor led to transverse relaxation time (T2) shortening compared to a blank control as shown by NMR relaxometry measurements. This process was also applied to the rapid and facile determination of concentrations of BPA in drinking water (tap water). Good linearity for all calibration curves was obtained, and the limit of detection (LOD) for BPA was 0.4 ng/mL in tap water.     

Electrochemical microdevice with separable electrode and antibody chips for simultaneous detection of pepsinogens 1 and 2

An electrochemical microdevice with separable electrode and antibody chips has been developed and applied to detect atrophic gastritis-related proteins, pepsinogen 1 (PG1) and pepsinogen 2 (PG2), based on sandwich-type enzyme-linked immunosorbent assays (ELISAs) with horseradish peroxidase (HRP)-labeled antibody. To fabricate the electrochemical device for simultaneous analysis of several proteins, the electrode chip with eight electrode elements was assembled along with an antibody chip with eight cavities containing immobilized anti-PG1 or anti-PG2. The immunoreactions occurring in the cavities of the device were detected simultaneously by amperometry. The labeled HRP in the cavity in the presence of hydrogen peroxide catalyzed the oxidation of ferrocenemethanol (FMA) to FMA+, which was detected electrochemically by the electrode chip. The amperometric responses of respective cavities in the device increased with increasing concentration of PG1 or PG2 of 0-50 ng/ml, ensuring the simultaneous detection of PG1 and PG2. The detection limits for both PG1 and PG2 were 0.6 ng/ml (S/N=2). The electrode chip was recovered easily by disassembling the electrochemical device; thereby, it was used repeatedly, whereas the antibody chip was discarded. No marked decrease in electrochemical responses was detected after repeated use. Reuse of the electrode chip is beneficial to reduce costs of protein analysis.     

Sensing HIV related protein using epitope imprinted hydrophilic polymer coated quartz crystal microbalance

We have developed a biomimetic sensor for the detection of human immunodeficiency virus type 1 (HIV-1) related protein (glycoprotein 41, gp41) based on epitope imprinting technique. gp41 is the transmembrane protein of HIV-1 and plays an important role in membrane fusion between viruses and infected cells. It is an important index for determining the extent of HIV-1 disease progression and the efficacy of therapeutic intervention. In this work, dopamine was used as the functional monomer and polymerized on the surface of quartz crystal microbalance (QCM) chip in the presence of template, a synthetic peptide with 35 amino acid residues, analogous to residues 579-613 of the gp41. This process resulted in grafting a hydrophilic molecularly imprinted polymer (MIP) film on the QCM chip. QCM measurement showed that the resulting MIP film not only had a great affinity towards the template peptide, but also could bind the corresponding gp41 protein specifically. The dissociation constant (K(d)) of MIP for the template peptide was calculated to be 3.17 nM through Scatchard analysis, which was similar to those of monoclonal antibodies. Direct detection of the gp41 was achieved quantitatively using the resulting MIP-based biomimetic sensor. The detection limit of gp41 was 2 ng/mL, which was comparable to the reported ELISA method. In addition, the practical analytical performance of the sensor was examined by evaluating the detection of gp41 in human urine samples with satisfactory results.     

Dual detection of COVID-19 antigens and antibodies using nanoscale fluorescent plasmonic substrates

There is a continuing need for biosensors that can be used in the diagnosis of COVID-19 infection and to measure a subject’s immune response to the virus itself (SARS-CoV-2). In this study, grating-coupled fluorescent plasmonic (GC-FP)-based detection of SARS-CoV-2 antigens was coupled with antibody detection to yield a dual-mode detection assay. Pairs of capture and detection antibodies were screened for direct detection of the SARS-CoV-2 nucleocapsid (Nuc) antigen, which were then combined with an existing GC-FP antibody detection assay. Nuc could be detected as low as 1 µg/mL concentrations, while antibodies were detectable to 50 ng/mL. The dual detection assay was tested by adding purified Nuc antigen to serum from a polymerase chain reaction (PCR)-positive COVID-19-infected individual. Using this sample, co-detection of Nuc antigen and anti-spike protein antibodies was successfully performed on a single GC-FP chip. Total assay time was 1 h, making this the first known example of rapid dual antibody and antigen detection on the same biosensor chip.     

Microfluidic electrochemical immunoarray for ultrasensitive detection of two cancer biomarker proteins in serum

A microfluidic electrochemical immunoassay system for multiplexed detection of protein cancer biomarkers was fabricated using a molded polydimethylsiloxane channel and routine machined parts interfaced with a pump and sample injector. Using off-line capture of analytes by heavily-enzyme-labeled 1 μm superparamagnetic particle (MP)-antibody bioconjugates and capture antibodies attached to an 8-electrode measuring chip, simultaneous detection of cancer biomarker proteins prostate specific antigen (PSA) and interleukin-6 (IL-6) in serum was achieved at sub-pg mL?1 levels. MPs were conjugated with ～90,000 antibodies and ～200,000 horseradish peroxidase (HRP) labels to provide efficient off-line capture and high sensitivity. Measuring electrodes feature a layer of 5 nm glutathione-decorated gold nanoparticles to attach antibodies that capture MP-analyte bioconjugates. Detection limits of 0.23 pg mL?1 for PSA and 0.30 pg mL?1 for IL-6 were obtained in diluted serum mixtures. PSA and IL-6 biomarkers were measured in serum of prostate cancer patients in total assay time 1.15 h and sensor array results gave excellent correlation with standard enzyme-linked immunosorbent assays (ELISA). These microfluidic immunosensors employing nanostructured surfaces and off-line analyte capture with heavily labeled paramagnetic particles hold great promise for accurate, sensitive multiplexed detection of diagnostic cancer biomarkers.     

Macro/nao-structured silicon as solid support for antibody arrays

To facilitate high-throughput biomarker discovery and high-density protein-chip array analyses of complex biological samples, a novel macro-and nanoporous silicon surface for protein microarrays was developed. The surface offers three-dimensional surface enlarging properties and spot confinement, enabling both high sensitivity bioassays and design of high density arrays. Reproducible manufacturing of the protein chip surface was accomplished as demonstrated by the low imprecision when standard IgG bioassays were performed at 100 pM antigen level on a series of protein chips scanned at widely different locations within a silicon wafer, as well as between different wafers from two different manufacturers. The relative standard deviation (RSD) of fluorescence spot intensity within an array on a chip was less than 20%. Mean spot intensity RSD was 19% for all 25 microarray chips in the study. Within-manufacturer-lot RSDs in chips from either manufacturer were <15% of mean spot intensity. The detection limit and dynamic range of the novel protein chip surface were examined to evaluate whether they match criteria required in a search for novel biomarkers such as for prostate cancer. Monoclonal IgG against prostate-specific antigen (PSA) was arrayed on the porous silicon chips. These were subsequently incubated in serum samples containing widely different levels of fluorescence-labeled PSA. Detection of PSA in serum at concentrations from 0.7 ng/mL (26 pM) up to 10⁴-fold higher levels verified assay characteristics required in the search for prostate biomarkers (e.g., kallikrein gene products) at clinically relevant levels.     

Are plasma insulin-like growth factor I (IGF-I) and IGF-binding protein 3 (IGFBP-3) useful markers of prostate cancer?

Increased concentrations of insulin-like growth factor I (IGF-I) and decreased insulin-like growth factor binding protein 3 (IGFBP-3) in serum have been proposed as markers of prostate cancer (CaP). The evidence for this, however, is contradictory. We assayed serum for IGF-I, IGFBP-3 and prostate-specific antigen (PSA) in patients with CaP and benign prostatic hyperplasia (BPH) and in healthy controls (HC). The mean +/- SD concentration of IGF-I in CaP (98.3 +/- 39.3 ng/mL; n = 15) was lower than in BPH (119 +/- 31.1 ng/mL; n=24) and HC (119 +/- 36.1 ng/mL; n=46), but the differences between the three groups were not statistically significant (p > 0.05). The mean IGFBP-3 concentrations in CaP (2691 +/- 1105 ng/mL; n = 16; p = 0.029) and BPH (2618 +/- 816 ng/mL; n = 26; p = 0.006) patients were significantly lower than that of the HC (3119 +/- 618 ng/mL; n=59), but the difference between the two groups of patients was not significant (p > 0.05). PSA concentrations in CaP (median = 80.8 ng/mL; n = 25) were significantly higher than those in BPH (median = 8.6 ng/mL; n = 39) (p < 0.001). Ninety-six percent of CaP and 72% of BPH patients had PSA concentrations >4.0 ng/mL; the proportions of patients with concentrations exceeding 20 ng/mL were 76% and 10%, respectively. We conclude that IGF-I and IGFBP-3 are inferior to PSA for CaP detection.     

Flow-injection resonance light scattering detection of proteins at the nanogram level.

A resonance light scattering (RLS) detection method for protein was developed, using a flow-injection system based on the enhancement of RLS signals from Biebrich scarlet (BS) by protein. The enhanced RLS intensities at 286.0 nm, in acidic aqueous medium, were proportional to the protein concentration over the range 0.005-18 microg/mL and 0.008-16 microg/mL for human serum albumin (HSA) and bovine serum albumin (BSA), respectively, with corresponding limits of detection (3sigma) of 5.00 ng/mL for HSA, and 7.80 ng/mL for BSA. The method was successfully applied to the quantification of total proteins in human serum samples.     

Label-free immunosensor for prostate-specific antigen based on single-walled carbon nanotube array-modified microelectrodes

We have fabricated a label-free electrochemical immunosensor using microelectrode arrays modified with single-walled carbon nanotubes (SWNTs). Label-free detection of a cancer marker, total prostate-specific antigen (T-PSA), was carried out using differential pulse voltammetry (DPV). The current signals, derived from the oxidation of tyrosine (Tyr), and tryptophan (Trp) residues, increased with the interaction between T-PSA on T-PSA-mAb covalently immobilized on SWNTs. The selectivity of our biosensor was challenged using bovine serum albumin (BSA) as the target protein. The detection limit for T-PSA was determined as 0.25 ng/mL. Since the cut-off limit of T-PSA between prostate hyperplasia and cancer is 4 ng/mL, the performance of our label-free electrochemical immunosensor seems promising for further clinical applications.     

基于纳米颗粒的赭曲霉毒素A高灵敏酶联免疫检测方法的建立及应用

赭曲霉毒素(ochratoxins)主要是由青霉菌Penicillium和曲霉菌Aspergillus产生的有毒次级代谢产物,常见于发霉或发酵的农产品中,其中赭曲霉毒素A(ochratoxin A,OTA)毒性最强且最为普遍。OTA是粮食作物和饲料的重要污染物,在加工、储存或运输过程中均可产生,具有肾毒性和免疫毒性,可通过蓄积作用发挥毒性效应,对人类和动物健康造成严重威胁。本研究通过将OTA单克隆抗体包被于纳米磁珠(magnetic nanoparticles,MNPs)表面,获得具有免疫活性的磁珠抗体复合物(MNPs-Anti OTA),并制备生物素标记的偶联抗原OTA-BSA-Bio,后续采用链酶亲和素标记的纳米金颗粒(Strep-HRP-AuNPs)催化底物进行信号检测,最终建立了OTA高灵敏检测方法(MNPs-bs-AuNPs-ELISA)。在最优条件下,经计算该方法检测下限(IC10)为0.01ng/mL,检测区间(IC20-IC80)为0.02-0.73ng/mL,半数抑制率(IC50)为0.13ng/mL。与OTA类似物OTB、OTC交叉反应性为4.3%和8.1%,对其他常见真菌毒素AFB1、ZEN、FB1、DON、CIT和PAT均无交叉反应。玉米、面粉和大豆样本中的加标回收率可达85.6%-115.7%,对天然样本中OTA含量的检测结果表明,该方法与LC-MS/MS相关性良好。本研究建立的MNPs-bs-AuNPs-ELISA可满足谷物及饲料样本中OTA的快速、高灵敏度定量检测,成本较低,具有很好的应用前景。     

Application of hollow fiber liquid phase microextraction coupled with high-performance liquid chromatography for the study of the osthole pharmacokinetics in cerebral ischemia hypoperfusion rat plasma

A simple and solvent-minimized sample preparation technique based on two-phase hollow fiber liquid phase microextraction has been developed and used to quantify the osthole in cerebral ischemia reperfusion rat plasma following oral administration. The analysis was performed by reversed phase high performance liquid chromatography with fluorescence detection. Extraction conditions such as solvent identity, agitation rate, salt concentration, extraction time, and length of the hollow fiber were optimized. Under the optimized conditions, the linear range of osthole in rat plasma was 5-500 ng mL(-1) (r(2) = 0.9997). The limit of detection (LOD) was 2 ng mL(-1) (S/N = 3) with limit of quantification (LOQ) being 5 ng mL(-1). The validated method has been successfully applied for pharmacokinetic studies of osthole from cerebral ischemia reperfusion rat plasma after oral administration.     

Preparation of a visual protein chip for detection of IgG against Treponema pallidum

A rapid, visual protein chip method for the detection of IgG against Treponema pallidum was developed using Staphylococcus protein A-modified gold nanoparticles. After recombinant T. pallidum antigen Tpp47 was arrayed on aldehyde-coated slides, qualitative and semiquantitative assay results were obtained from the hybridization signal and the subsequent gray stain values. The Tpp47-specific serum antibody was bound to immobilized Tpp47 antigen on the functionalized glass slides. The Staphylococcus protein A-modified gold nanoparticle probe exclusively recognized anti-Tpp47 IgG. The "sandwich" format hybridization signal was then amplified with silver staining, a high sensitivity visual detection technique. It took 3.5 h to prepare the detection protein chip, but only 20 min for real sample detection. The lowest titer of detectable antibody for this method was 1:128, which correlates to approx 1 ng/mL. Furthermore, the results can only be seen unaided, which drastically reduces the cost of detection, but can also be kept for a long time. The data also confirmed the proposed method's specificity, sensitivity, and convenience. As a result, the method could be applied as an alternative diagnostic tool in the clinical diagnosis of T. pallidum.     

Simultaneous and rapid detection of six different mycotoxins using an immunochip

Mycotoxins are highly toxic contaminants in food, animal feed, and commodities. The study has developed an immunochip for quantifying the concentrations of six mycotoxins: aflatoxin B1, aflatoxin M1, deoxynivalenol, ochratoxin A, T-2 toxin, and zearalenone, which were added to drinking water. The complete antigens (Ags) of the mycotoxins were contact printed and immobilized onto agarose-modified glass slides with 12 physically isolated subarrays, based on the reaction of both diffusion and covalent bond. The optimal concentration of each antigen and antibody (Ab) was obtained using an Ag-Ab immunoassay. Based on the indirect competitive immunoassay for the simultaneous detection of six mycotoxins in one single chip, six standard curves with good logistic correlation (R(2)>0.97) were respectively plotted. The working ranges (0.04-1.69, 0.45-3.90, 20.20-69.23, 35.68-363.18, 0.11-1.81, and 0.08-7.47 ng/mL, respectively) were calculated, as well as the median inhibitory concentrations (0.31±0.04, 1.49±0.21, 34.54±1.30, 134.06±11.75, 0.49±0.05, and 1.54±0.22 ng/mL, respectively), when six mycotoxins were detected simultaneously. Finally, the recovery rates in drinking water generally ranged from 80% to 120% on the same chip, with an intra-assay coefficient of variation lower than 15%. We successfully established an immunochip for simultaneous detection of six mycotoxins within 4h, with advantages of using minimal samples and being visually semiquantitative with our naked eyes. In summary, the method could be developed on one single chip for detecting multiple contaminants in actual samples.     

Chemiluminescence determination of moxifloxacin in pharmaceutical and biological samples based on its enhancing effect of the luminol–ferricyanide system using a microfluidic chip

A sensitive determination of a synthetic fluoroquinolone antibacterial agent, moxifloxacin (MOX), by an enhanced chemiluminescence (CL) method using a microfluidic chip is described. The microfluidic chip was fabricated by a soft‐lithographic procedure using polydimethyl siloxane (PDMS). The fabricated PDMS microfluidic chip had three‐inlet microchannels for introducing the sample, chemiluminescent reagent and oxidant, and a 500 µm wide, 250 µm deep and 82 mm long microchannel. An enhanced CL system, luminol–ferricyanide, was adopted to analyze the MOX concentration in a sample solution. CL light was emitted continuously after mixing luminol and ferricyanide in the presence of MOX on the PDMS microfluidic chip. The amount of MOX in the luminol–ferricyanide system influenced the intensity of the CL light. The linear range of MOX concentration was 0.14–55.0 ng/mL with a correlation coefficient of 0.9992. The limit of detection (LOD) and limit of quantification (LOQ) were 0.06 and 0.2 ng/mL respectively. The presented method afforded good reproducibility, with a relative standard deviation (RSD) of 1.05% for 10 ng/mL of MOX, and has been successfully applied for the determination of MOX in pharmaceutical and biological samples. Copyright © 2013 John Wiley & Sons, Ltd.     

Prognostic value of cytokeratin 19 fragment (CYFRA 21-1) and cytokeratin-positive cells in bone marrow samples of breast cancer patients

The aim of this study was to investigate the relationship between the detection of micrometastatic cells by immunocytochemistry (ICC) with an anticytokeratin antibody and cytokeratin fragment (CYFRA 21-1) expression detected by an immunofluorescent assay in bone marrow of breast cancer patients. Micrometastatic CK+ cells were screened with a pancytokeratin antibody A45 B/B3 from bone marrow aspiration samples of 102 breast cancer patients (65 primary tumors, 10 local recurrences and 27 distant metastases). CYFRA 21-1 levels were assessed in bone marrow supernatant of these patients before collection of the mononucleated interface cells on a Ficoll-Hypaque density gradient and in 20 control patients. CYFRA 21-1 and CK+ cell detection by ICC were both correlated with clinical stage. CYFRA 21-1 was significantly elevated in patients with micrometastatic disease detected by ICC: 4.77 ng/mL (+/- 10.87 SD) versus 1.00 ng/mL (+/-1.36 SD) in patients with negative ICC (p=0.01). In univariate analysis, a CYFRA 21-1 value > or =1 ng/mL and the presence of CK+ cells were associated with a poorer survival for patients with stage I to III breast cancer (n=65). On multivariate analysis, only pathological nodal status and presence of CK+ cells in bone marrow were independent prognostic factors for overall survival. In conclusion, in this series CYFRA 21-1 was correlated with detection of CK+ cells by ICC in bone marrow, but cannot replace ICC. The presence of CK+ cells in bone marrow remains a strong independent prognostic factor in primary breast cancer.     

A simple electrochemical aptasensor for ultrasensitive protein detection using cyclic target-induced primer extension

A simple electrochemical aptasensor was developed for ultrasensitive protein detection by combining a novel strategy of cyclic target-induced primer extension (CTIPE) with an aptamer-hairpin probe and enzyme-amplified electrochemical readout. In the presence of protein target, the immobilized aptamer-hairpin probe recognized the protein to trigger primer extension reaction by target-induced conformational transition, which released the protein from replicated DNA duplex. The released target could cyclically bind with other aptamer-hairpin probes and trigger new primer extension, leading to formation of numerous biotin-tagged DNA duplex, which significantly amplified the protein recognition event and facilitated the subsequent enzymatic signal enhancement, leading to an ultrasensitive electrochemical aptasensor. Using human vascular endothelial growth factor as a model protein, the designed aptasensor could detect protein down to 0.82 pg mL(-1) with a linear range from 1 pg mL(-1) to 1 ng mL(-1). The proposed aptasensor was amenable to quantification of protein in complex biological matrixes, and would become a simple and powerful tool for bioanalysis and clinic diagnostic application.     

Biocompatible in-tube solid-phase microextraction coupled to HPLC for the determination of angiotensin II receptor antagonists in human plasma and urine

A poly (methacrylic acid-ethylene glycol dimethacrylate, MAA-EGDMA) monolithic capillary was used for the in-tube solid-phase microextraction (in-tube SPME) of several angiotensin II receptor antagonists (ARA-IIs) from human plasma and urine. Under the optimized extraction condition, the protein component of the biological sample was flushed through the monolithic capillary, while the analytes were successfully trapped. Coupled to HPLC with fluorescence detection, this on-line in-tube SPME method was successfully applied for the determination of candesartan, losartan, irbesartan, valsartan, telmisartan, and their detection limits were found to be 0.1-15.3ng/mL and 0.1-15.2ng/mL in human plasma and urine, respectively. The method was linear over the range of 0.5-200ng/mL for telmisartan, 5-2000ng/mL for candesartan and irbesartan, 10-2000ng/mL for valsartan, and 50-5000ng/mL for losartan with correlation coefficients being above 0.9985 in plasma sample and above 0.9994 in urine sample. The method reproducibility was evaluated at three concentration levels, resulting in the R.S.D. <7%. The poly (MAA-EGDMA) monolithic capillary was demonstrated to be robust and biocompatible by using direct injections of biological samples.     

PSA在前列腺增生和前列腺癌中的诊断价值

目的：探讨临床上检测前列腺特异性抗原（PSA）的变化情况对前列腺增生和前列腺癌等疾病的诊断价值。方法：采用回顾性分析的方法,选取2010年6月至2012年4月在我院泌尿科接受治疗的前列腺增生患者64例定义为前列腺增生组（BPH）,前列腺癌患者83例定义为前列腺癌组（PCa）,另选取同期接受体检的健康人群137例作为对照组。分别检测三组患者入院时的游离前列腺特异性抗原和总前列腺特异性抗原的水平变化情况。对比并分析三组检测结果。结果：经检测,前列腺增生患者的血清总PSA明显高于对照组健康人群的正常值,而前列腺癌患者的血清总PSA比前列腺增生患者增高的更为明显。对照组游离PSA为（2．78±0．94）ng／mL,总PSA（1．05±0．57）ng／mL,游离PSA与总PSA的比值为,（0．38±0．61）;前列腺增生患者游离PSA为（6．36＋3．24）ng／mL,总PSA为（1．64±0．76）ng／mL,游离PSA与总PSA的比值为（0．26±0．23）;前列腺癌患者游离PSA为（12．42±4．97）ng／mL,总PSA为（1．44±0．78）ng／mL,游离PSA与总PSA的比值为（0．12±0．16）。组间比较差异明显,具有统计学意义（P〈0．05）。结论：对患者的PSA进行检测,对前列腺增生和前列腺癌的诊断具有良好的辅助作用和,临床价值。