1.Introduction

Metallic biomaterials are used in various medical applications,mainly due to their attributes of high strength,fracture resistance,good electrical conductivity,and biocompatibility[1].Surface modification of biomaterials is considered to be an effective method to control cell adhesion,migration,orientation,shape,and even gene expression in in vitro and in vivo studies[2-6].For example,good adhesion between cells and implants is crucial for fracture repair,as it affects not only the growth and metabolism of bone tissue and physiology,but also the repair and reconstruction of bone tissue under conditions of injury[7-9].Compared with conventional surface-modification techniques such as photolithography,electrospinning,electron beam lithography,and chemical patterning[10-12],the main advantages of laser techniques are the lack of contaminants,high enhancement factor,low cost with high uniformity and reproducibility,and capability to tailor topography at the microscale or even nanoscale.Therefore,laser microprocessing has attracted considerable attention from both scientists and engineers in recent years.Martínez-Calderon et al.[13]investigated the culturing capabilities of laser-induced periodic surface structures(LIPSS)on stainless steel,and found that cells preferentially align and attach to the LIPSS with particular direction.Cunha et al.[14]reported that the stretching of human mesenchymal stem cells(hMSCs)could be induced by LIPSS for orthopedic implants via ultrafast laser surface texturing.Dumas et al.[15]fabricated three types of biomimetic patterns on the surface of a titanium(Ti)alloy,thereby showing that the combination of LIPSS and micro pits could enhance osteogenic potential.As a promising biomaterial material with a degradation behavior and elastic modulus similar to human bones,magnesium(Mg)alloys have attracted a great deal of attention for use as implants[16-18].However,high ion release and a tremendous change of local pH in the rapid degradation at the Mg alloy surface result in a survival environment that is quite severe for cells and that greatly reduces cell viability[12].The improvement of commercial Mg alloys through corrosion resistance using laser surface modification was subsequently proposed,and has been widely studied in the past decades[19,20].To the best of our knowledge,cell behavior at the surfaces of laser-treated Mg alloys has not been reported in the public domain.

(3)必须保证水封排气内水面的高度。水位过高，炉内压力增加，水封有可能不能顺利排气。水位过低，水封密封效果降低，一旦无水，后果非常严重。

基于13个形态数据的聚类分析，将106份样本分成了8个组(图2，表4)。方差分析表明，除中肋长度外的12个性状均存在组间极显著差异(P < 0.001)，同时与地理来源有较强对应性。

Moreover,liquid biopsies play an increasingly important role in the diagnosis and treatment of cancer due to their noninvasiveness,convenient sampling,and capability for real-time monitoring in the detection of cells and DNA[21-24].Detection indicators with excellent specificity,sensitivity,and stability are very necessary in order to improve the accuracy and efficiency of lipid biopsies.Surface-enhanced Raman scattering(SERS)has been widely used for biomaterial analyses and the detection of lowconcentration biochemical molecules based on vibrational spectroscopic information of molecules obtained by inelastic scattering with photons.In recent years,the periodic surface structures of detected substrates have been successfully employed to enhance Raman scattering,resulting in sensitive detection and efficient remediation.A variety of laser-induced SERS methods have been reported,most of which can be simply classified into two categories:silicon(Si)substrates and non-Si substrates.In regard to Si substrates,Xu et al.[25]found that micro-and nanostructures with particle sizes below 100 nm at the Si surface provided the greatest improvement in Raman scattering.Zhu et al.[26]reported that SERS enhancement factors as high as 3×107 had been achieved on laser-ablated gold(Au)-coated Si wafers with three-dimensional(3D)micro-or nano-structures for Rhodamine 6G(R6G)molecules,as a result of the enhanced surface area and Au nanoparticles with different sizes.Parmar et al.[27]fabricated nano-scaffolds decorated with metal nanoparticles on black Si substrates,showing that surface plasmonic effects and‘‘hot spots”at laser-textured surfaces could lead to significant enhancement of the Raman signal.In regard to non-Si substrates,Buividas et al.[28]demonstrated a direct laser write approach to create nano-textured surfaces on sapphire;these surfaces showed a SERS signal up to 15 times higher with a signal uniformity two times better than commercial SERS-sensing substrates.Rebollar et al.[29]fabricated Au-coated LIPSS on spin-coated polymer thin films by means of a nanosecond laser,and investigated their characterization as substrates for SERS.The enhancement factor for the Au-coated substrates was estimated to be eight orders of magnitude,while an additional enhancement of around ten was mediated by the presence of LIPSS.Unfortunately,no reports on biocompatible substrates are available in the public domain.

The present work investigates the feasibility of laser microprocessing for biomedical metallic materials including Mg and Ti alloys,with potential applications in cell adhesion and liquid biopsy.Laser surface modification of a magnesium-gadoliniumgallium (Mg-Gd-Ga) alloy is examined first.Next,the biocompatibility—that is,cell viability and adhesion—of the as-received and laser-modified surfaces is evaluated using direct cell culture.We then examine the laser surface modification of a Mg-Gd-Ca alloy and apply the direct cell culture method to investigate initial cell viability and adhesion in order to compare the biocompatibility of the as-received and laser-modified surfaces.In addition,we investigate the capability of a femtosecond laser to achieve 3D LIPSS on a Ti alloy in order to enhance SERS measurement by enlarging the effective signal of the detected samples in order to simultaneously improve both efficiency and sensitivity.

2.Laser melting and nanostructuring of Mg alloys for biocompatibility enhancement

As a promising alternative to commercial Mg alloys,Mg-Gdbased alloys show a high performance in terms of biocompatibility and qualified mechanical properties.Like certain other rare earth elements,Gd has good biocompatibility.In addition,due to its large solubility in Mg at the eutectic temperature and intermetallic phase formation,introducing Gd into an alloy is beneficial in order to strengthen the formation of a solid solution and precipitation.

按国家法律规定是让集体内部的人先流转，集体所在的人不流转的情况下才让外人流转。比如乙村的地先由乙村人流转，村里人不流转才接着让其他人流转。对我来说我出生在甲村，后来我家在乙村也有宅基地。这样我虽然不是农村户口，不在农村居住，但是我家在村里有地的话起码也能算上是这村的后人。……要是在其他乡镇，有农民提出来说“他就不是这村的人，凭什么包村里的地？我还想包地呢！”这时候土地流转可能就进行不下去了。现在最起码我有这村里的地，算这个集体的人。(宋某访谈资料)

2.1.Experimental procedures

2.1.1.Materials

As a solid solution element in a Mg matrix and in the Mg-Gd eutectic phase of a Mg-6Gd-based alloy,calcium(Ca)is helpful to refine the grain and improve the yield strength of the alloy.Hence,hot-extruded Mg-6Gd-0.6Ca alloy bars were chosen in this case.The bar diameter was 9 mm,and a 4 mm slice was ground using silicon carbide(SiC)paper.After being washed with ethanol,the alloy was etched in a solution of 0.5%picric acid(1 g picric acid,24 mL distilled water,24 mL acetic acid,and 200 mL ethanol).

As shown in Eq.(1),Mg reacts with water(H2O)to form hydrogen gas(H2).The quantity of H2 evolution is not in fluenced by the corrosion products formed at the surface of the Mg.The H2 evolution rate,νH(mL·cm-2·d-1),was estimated by measuring the total H2 released over a certain period of time[36].Table 2 gives the total H2 release and the H2 evolution rate which corresponds to the average corrosion rate.

Fluorescence investigation of the cell adhesions showed that cells at the laser-melted-and-LIPSS surfaces(Fig.7(b))presented an elongated shape with a smaller surface area than those at the laser-melted surfaces(Fig.7(a)).This is mainly due to cell shape anisotropy during cytoskeleton reorganization caused by LIPSS,which can act as an anisotropic and persistent mechanical stimulus[17].Cell density at the laser-melted surfaces was higher than at the laser-melted-and-LIPSS surfaces,due to the combined effect of corrosion resistance and surface topography.On the one hand,biocorrosion tests showed that the corrosion resistance of the laser-melted surface was better than that of the laser-meltedand-LIPSS surface,resulting in a lower pH value and Mg2+concentration in the environment,which may better permit cells to survive and adhere.On the other hand,it has been reported that a surface structure with dimensions exceeding 70 nm may affect the integrin clustering efficiency,leading to a reduction of cell adhesion[20].In this work,LIPSS with a period of 900 nm and depth of 250 nm were shown to reduce cell adhesion.

The Mg-6Gd-0.6Ca alloy surface was modified under argon(Ar)gas protection by a continuous-wave(CW) fiber laser centered at 1064 nm with a spot size of 100μm.The scanning speed and the power density were respectively set at 70 mm·s-1 and 2.04 × 106 W·cm-2.

Surface texturing was carried out on the laser-melted surface using a Ti:sapphire chirped-pulse regenerative amplification laser system(with the center wavelength of 1064 nm,the pulse width of 800 fs,and the repetition rate of 400 kHz)at normal incidence in air at ambient temperature and pressure.The diameter of the Gaussian beam was measured to be about 35μm.LIPSS were produced by adjusting the laser parameters.

2.1.3.Surface characterization

处置程序：①现场停产撤人；②安排矿、科级人员到调度台指挥、副科级以上人员到现场；③安监处安排科长在调度台指挥、副科以上人员现场监护安全；④按事故类型由分管矿领导或值班矿领导在调度台指挥，根据需要副总以上矿领导到现场，见图1。

2.1.4.Biocorrosion tests

The degradation rate of the Mg alloy was monitored by the evolved hydrogen volume.The hydrogen volume measurements of the as-received,laser-melted,and laser-melted-and-LIPSS specimens were studied in Hank's balanced salt solution(HBSS)for 34 h at 37°C in a circulating water bath.The HBSS's volume per unit area of sample was 20 mL·cm-2.Fig.1 shows the testing setup,and details of the measurement setup can be found in the literature[30].The hydrogen evolution volume was measured every 2 h.

水生产力与人水关系的辩证关系，是马克思关于生产力与生产关系的基本哲学原理在水资源利用与管理中的具体体现。按照生产工具的发展水平，水生产力的发展可分为人工、机械化、电气信息化、社会化及和谐化等5个阶段，分别对应于人水关系的原始和谐、自由供水、总量控制、反哺自然及人水和谐等5个阶段。不同的人水关系发展阶段必须有相应的水生产力水平支撑，否则人水关系及相应政策将仅是理念，难以真正实施。我国正在实施的最严格水资源管理制度属于总量控制阶段，处在人水关系发展演变的关键时期。

Fig.1.Setup of hydrogen evolution measurement during immersion measurement.

The electrochemical behavior was measured at 37°C in HBSS using an electrochemical workstation(CHI660e,CH Instruments Inc.,USA).The chemical composition of the HBSS solution is provided in Table 1.A three-electrode system was employed with a saturated calomel electrode as the reference electrode and a carbon electrode as the counter electrode;the specimen was the working electrode,with an exposed surface area of 0.64 cm2.Tafel curves of the as-received and laser-modified specimens were measured at a scanning rate of 1 mV·s-1.

2.1.5.Cell culture

Murine calvarial preosteoblasts(MC3T3-E1,Peking Union Medical College,China)were applied to examine the in vitro biocompatibility of the Mg alloy.The MC3T3-E1 were cultured in 10%fetal bovine serum(FBS),Dulbecco's modified Eagle's medium(DMEM),100 μg·mL-1 streptomycin at 37 °C,and 100 U·mL-1 penicillin in a humidified atmosphere of 5%carbon dioxide(CO2).

Surface topography was measured and characterized by optical spectroscope(LV150N,Nikon Corporation,Japan)and scanning electron microscope(SEM)with an energy dispersive spectrometer(SU8010,Hitachi Ltd.,Japan).The phase was confirmed using an X-ray diffractometer(D/max2200PC,Rigaku Corporation,Japan).

The Ti6Al4V substrate was cut into square samples with dimensions of 10 mm×10 mm×1 mm.Prior to laser processing,these samples were polished with SiC sandpaper in different grades of roughness(from 380 to 4000,in order to obtain a surface roughness less than 0.01μm)and subsequently ultrasonically cleaned in acetone.

The specimens werefirst sterilized by exposure to ultraviolet light radiation for 2 h.Cells were seeded by adding 70 mL of MC3T3-E1 cell suspension onto the surface,and were then cultured for 48 h.The initial density was 7×104 per specimen placed into a 24 well plate.For the cell morphology measurement,the cell-seeded scaffolds were cleaned three times with phosphate buffer solution(PBS)and fixed in paraformaldehyde for 30 min,while the specimens were rewashed three times using PBS and then dehydrated using graded ethanol.After vacuum drying and spraying with gold particles,the specimens were characterizedusing SEM.For the fluorescence microscopy investigation,the specimens were soaked in 70%ethanol solution for cell fixation,and cell components were colored using 5 mg·L-1 acridine orange dye.The fluorescence microscopy results were obtained by a fluorescence microscope(IX71,Olympus Corporation,Japan)equipped with a charge-coupled device(CCD)camera(iXON EM+DU-897ECSO-UVB,Andor Technology Ltd.,Ireland).

Table 1 Chemical composition of HBSS.

Composition Concentration(mmol·L-1)Na+ 142.0 K+ 5.8 Mg2+ 0.8 Ca2+ 2.5 Cl- 145.0 HCO-4.2 H2PO-3 0.4 HPO2-4 0.3 SO2-4 4 0.8

2.2.Results and discussion

2.2.1.Morphology and microstructure

As shown in Fig.2(a),the as-received Mg alloy consisted of an α-Mg matrix and β-phase Mg5Gd[31,32].After laser modification,the X-ray diffraction(XRD)results revealed that the laser-melted surface contained only theα-Mg phase;no Mg-Ca eutectic phase was found,either in the substrate or in the melted layer.This was attributed to the low Ca content in the Mg-6Gd-0.6Ca alloy[33].Fig.2(b)shows the typical solidification microstructure and surface morphology of the Mg-6Gd-0.6Ca after laser modification.A 400μm thick melted layer with a solidification microstructure was generated,and the main content comprised small columnar grains of theα-Mg phase[34].During laser surface processing,the temperature of the substrate increases dramatically when irradiated by the laser beam,and instantaneously reaches the melting point.When the laser beam moves on,the temperature drops immediately,leading to a huge temperature gradient and a rapid cooling rate in the melted layer.Consequently,no β-phase was found in the laser-melted layer.The surface topography of the LIPSS generated on the laser-melted surface is shown in Fig.3(a).According to the surface profile illustrated in Fig.3(b),the depth and period of the LIPSS are around 250 and 900 nm,respectively.

2.2.2.Hydrogen evolution

Fig.4 presents the hydrogen evolution data for the as-received,laser-melted,and laser-melted-and-LIPSS samples during immersion in HBSS.The Mg corrosion reaction equation is provided below[35]:

洒布材料选用AH—70号热沥青，用量为1.6kg/m2，然后再洒布一层经拌和站加热除尘的石灰岩石屑，其厚度为0.5cm，粒径为0.5～1.0cm，其中掺加0.4%沥青。下封层洒布采用智能碎石洒布车及智能沥青洒布车进行洒布。并用胶轮压路机进行碾压。

Fig.2.Microstructure and phase compositions of Mg alloy before and after laser treatment.(a)XRD results;(b)cross-sectional view of laser-melted Mg alloy.

Fig.3.(a)SEM and(b)atomic force microscopy(AFM)topography of LIPSS on the laser-melted Mg alloy surface.

Fig.4.Volume of hydrogen generated on as-received,laser-melted,and lasermelted-and-LIPSS surfaces during immersion in HBSS for 48 h.

3.1.2.Laser processing

Table 2 The total H2 release and the H2 evolution rate.

Specimens Total H2 release(mL)H2 evolution rate(mL·cm-2·d-1)As-received 205.0 25.625 Laser-melted 7.8 0.975 Laser-melted-and-LIPSS 16.6 2.075

2.2.3.Electrochemical measurement

到了黑夜，王宝生就来到2楼的瞭望台，透明的玻璃墙壁把呼啸的寒风隔离开，他独自一人站在这个山头上，冷了就搓搓手、哈哈气，无聊了就来回踱步，默默地瞭望着万亩大山，想想那孤寂之感就让人喘不过气。

Fig.5.Potentiodynamic polarization results of as-received,laser-melted,and lasermelted-and-LIPSS specimens in HBSS.

Table 3 E corr and i corr for as-received,laser-melted,and laser-melted-and-LIPSS in HBSS.

Specimens E corr(V) i corr(mA·cm-2)As-received -1.508 0.05540 Laser-melted -1.406 0.00158 Laser-melted-and-LIPSS -1.421 0.00484

As shown in Fig.5,a polarization study of the as-received,lasermelted,and laser-melted-and-LIPSS specimens was performed in HBSS.Table 3 summarizes their corrosion potential(E corr)and corrosion current density(i corr).The E corr value of the laser-melted specimen exhibited a less-negative value(-1.406 V)than that of the as-received specimen(-1.508 V).After LIPSS were produced on the melted surface,the E corr shifted slightly to a more negative value(-1.421 V),although this value was still less negative than that of the as-received microstructures.This finding indicates increased cathodic behavior compared with the melted microstructures,but decreased cathodic behavior compared with the as-received microstructures,mainly because the LIPSS induced on the melted surface may destroy the surface and reduce the thickness of the melted layer.This finding agrees well with the immersion test results shown in Fig.4.Furthermore,the i corr of the laser-melted-and-LIPSS specimen was much lower than that of the as-received specimen,but slightly higher than that of the laser-melted specimen.This indicates that the corrosion rate of the laser-melted-and-LIPSS specimen is lower than that of the as-received specimen but higher than that of the laser-melted specimen.

水是基础性自然资源和战略性经济资源。水利是国民经济和社会发展的重要基础设施和基础产业。在新的发展阶段，如何更好地发挥水利行业的支撑和保障作用，是摆在我们面前重大而紧迫的课题。

2.2.4.In vitro biocompatibility

As shown in Fig.6,cell spreading behaviors including distribution and morphology were investigated at the surfaces of the asreceived,laser-melted,and laser-melted-and-LIPSS specimens for 48 h after cell seeding.Due to poor corrosion resistance,no cell was observed on the as-received surface.After heavy corrosion took place during culturing,the formation of rod-like crystals was observed at the surface,as shown in Fig.6(a).The viability of MC3T3-E1 cells is affected by the increased pH value and large release of Mg2+ions caused by corrosion[38].In contrast,cells exhibited good adhesion behavior and spread anisotropically on all laser-melted surfaces,as shown in Fig.6(b)and 6(c).Fig.6(c)indicates that the cell spreading was anisotropic on the lasermelted-and-LIPSS surfaces,with a large number of focal adhesions being presented as filopodia.Since filopodia usually act as nanosensors that allow cells to explore the external environment,the morphology of the filopodia indicates cell migration.High magnification of the focal adhesions in Fig.6(d)reveals that the cell is migrating along the direction of the LIPSS,with a tree-like structure in the front and back areas.

2.1.2.Laser processing

基于灰靶理论的“一带一路”沿线省份科技创新能力评价…………………………………………冯俊华 张庆妮 徐青青（3.25）

Therefore,we propose that the LIPSS at the melted surface is able to promote the growth of osteoblasts and accelerate ossification due to good adhesion;this can induce the cells that participate in fracture healing to gather at the end of a fracture and accelerate the healing of the fracture.

3.Large-scale nanostructuring of Ti alloys for SERS

Fig.6.SEM images of osteoblast shapes after cultured for 48 h on(a)as-received,(b)laser-melted,and(c)laser-melted-and-LIPSS surfaces;(d)zoomed-in area of cell protrusion,shown as the black box in(c).

Fig.7.Fluorescence images of osteoblast shapes on(a)laser-melted and(b)laser-melted-and-LIPSS surfaces after culturing for 48 h.

Although substrates have recently been designed to allow sensitive detection in the form of a specific Raman fingerprint via SERS,it is a challenging task to devise a simple approach for fabricating an SERS-responsive substrate based on biomedical materials and having biocompatibility.In this work,LIPSS were created on the surface of a bio-titanium alloy using laser irradiation with a femtosecond laser.To the best of our knowledge,this is the first report that realizes the capability of LIPSS to provide an enhanced spectroscopic signal via SERS,with the potential for application in the fields of bio-detection and bioimaging.

（2）草海湿地沉积物重金属元素除Cu、As平均含量未超过贵州省土壤元素背景值，其他5种重金属均超出贵州省土壤背景值含量，尤其是Cd，高达背景值21倍，可见草海存在严重的Cd污染。根据潜在生态危害指数的综合分析可知：Cr、As、Pb和Zn污染较严重，其中Cd和Hg污染最严重，已达到极强的潜在生态危害程度。

3.1.Experimental procedures

3.1.1.Materials

2.1.6.Cell viability and cell morphology

第2个数据集为SisFall,该数据集使用可穿戴嵌入式设备采集志愿者运动过程中的三轴加速度计、陀螺仪数据,采样频率同样为200 Hz,设备被固定在志愿者的腰部。38名志愿者被分为两组:23名年轻志愿者及1名60岁的柔道运动员提供了15种跌倒姿势和19种非跌倒运动数据;其余14名老年志愿者只提供15种非跌倒运动数据。利用时间窗对数据集进行序列化预处理后,我们共得到3 753例非跌倒数据和1 798例跌倒数据。

As shown in Fig.4,the hydrogen volumes of the as-received,laser-melted,and melted-and-LIPSS specimens were 205,7.8,and 16.6 mL,respectively,after 48 h immersion.This result indicates that both the laser-melted and the laser-melted-and-LIPSS specimens can effectively improve the corrosion resistance of the Mg-6Gd-0.6Ca alloy and significantly reduce the degradation rate in HBSS solution.The corrosion rate of the melted-and-LIPSS specimens,as estimated from the rate of H2 evolution,was slightlyhigher than that of the melted specimens and was about 12 times slower than that of the as-received specimen.This may be due to the β-Mg5Gd phase in the as-received specimen,which may act as a galvanic cathode and accelerate matrix corrosion[34].After laser surface modification,the dissolution of the β phase into the matrix significantly reduced the amount of galvanic corrosion.Moreover,the LIPSS induced on the melted surface may increase the surface area and reduce the thickness of the melted layer,resulting in an increased corrosion rate compared with the lasermelted specimens.As shown in Table 2,the H2 evolution rates of the as-received,melted,and melted-and-LIPSS specimens were 25.625,0.975,and 2.075 mL·cm-2·d-1,respectively,after 48 h immersion.It is known that the H2 adsorption rate tolerated by the human body is 2.25 mL·cm-2·d-1[37].Thus,the H2 evolution rates of both the laser-melted and the laser-melted-and-LIPSS specimens were lower than the tolerance value of a human body.

As shown in Fig.8,LIPSS were fabricated by a Ti:sapphire laser system(with the center wavelength of 800 nm,the pulse width of 35 fs,and the repetition rate of 400 kHz)at normal incidence in air at ambient temperature and pressure.The diameter of the Gaussian beam was measured to be 35μm.An electron beam evaporator was used to deposit gold(99.99%pure gold)thin film on the laser-fabricated Ti6Al4V surfaces for SERS.

3.1.3.Surface characterization

The surface topography was studied by SEM(SU8010,Hitachi Ltd.,Japan)and atomic force microscopy(AFM;ICON,Bruker Corporation,USA)in contact mode.Nanoscope Analysis 1.4 software was applied to visualize and study the 3D AFM scans on 10μm×10μm areas.

3.1.4.SERS measurement

Crystal violet(CV;≥98%purity,Sigma-Aldrich)was applied as a probe molecule.A solution drop with a volume of about 5μL was dropped onto the substrate and then dried in air.The dried substrate was used to carry out SERS signal measurement via Raman spectroscopy.The Raman microscope system(HORIBA Jobin Yvon,France)used to record the Raman spectra was equipped with a diode laser with a 532 nm excitation source and a CCD operating at a power level of around 5 mW.The acquisition time and accumulations were 10 s and 10,respectively.A 20×(numerical aperture=0.45)microscope objective lens and a thermoelectrically cooled CCD array with a 600 g·mm-1 diffraction grating were used to detect the signals.

在现代汉语中，“本”有很多意思，其中一个就是“自己这方面的”。生活中，我们可能会听到或看到“本位主义”这个词，它指在处理整体利益与个人利益之间的关系时，只顾“自己这方面的”利益，而不顾他人、整体的利益，对其他人或群体漠不关心。从道德的观点上来说，本位主义是自私的一种表现，通常是以牺牲他人、集体的利益而达到自己的目的。这是一种错误的心态，缺乏大局和长远意识，最终的结果是损害自己的利益。

3.2.Results and discussion

3.2.1.Morphology evolution

When the incident laser fluence was close to the ablation threshold of titanium[39],3D Au-coated LIPSS on a Ti6Al4V surface was fabricated.The 45°view of 3D LIPSS and the magnified view of the groove are respectively showed in Fig.9(a)and 9(b).Periodic structure with periodicity of 600 nm on the groove and nanoparticles with diameters ranging from 60 to 200 nm randomly distributed on the above structure can be obviously observed.Besides,on a larger scale of 3D LIPSS,the average periodicity of 30μm and the depth of 14μm were measured using optical spectroscope as shown in Fig.9(c).Furthermore,the cross-section profiles of the 3D LIPSS were also measured to confirm the results as shown in Fig.9(d).

Fig.8.(a)Experimental setup;(b)fabrication procedures of Au-coated 3D LIPSS.

Fig.9.Surface morphologies of 3D LIPSS fabricated on a Ti6Al4V surface.(a)SEM image;(b)magnified view of 3D LIPSS;(c)3D optical image of the 3D LIPSS;(d)cross-section profiles of the 3D LIPSS.

Fig.10.The SERS spectra of 10-3 mol·L-1 CV molecules adsorbed on the as-received surface,Au-coated as-received surface,3D LIPSS surface,and Au-coated 3D LIPSS surface.

3.2.2.SERS measurement

Fig.10 shows the Raman spectrum of 10-3 mol·L-1 diluted CV on different dried substrates.The CV spectrum includes the characteristic bands that are attributed to the corresponding molecular vibrations[40,41].For the as-received surface without coating,no Raman signal was observed.For the Au-coated as-received substrate,Raman signals with bands at 913,1617,and 1646 cm-1 were obtained.This observation indicates that gold nanoparticles are efficient for SERS,which is in agreement with previous findings[42].For the 3D LIPSS substrate,an intensity enhancement by a factor of 103 was observed,suggesting that 3D LIPSS are effective for the preparation of a SERS substrate.In regard to the Au-coated 3DLIPSS substrate,the sensitivity and intensity of the substrate were further enhanced after gold film deposition,since gold supports plasmons in the visible region[43].Table 4 lists the corresponding Raman signal enhancement factors(EFs)of the 3D LIPSS and Aucoated 3D LIPSS for the bands at 913,1176,1374,and 1618 cm-1.The EFs are the averaged values of five different locations.The results indicate that the highest EF reaches 6.7×103;this occurs at the Raman peak at 1374 cm-1 for the Au-coated 3D LIPSS.

Table 4 Enhancement factors for the 3D LIPSS SERS substrate.

Raman bands(cm-1) Au-coated 3D LIPSS 3D LIPSS 913 2132 538 1176 2111 319 1374 6732 1074 1618 1123 279

Local surface plasmon resonance(LSPR)and surface plasmon polaritons(SPPs)will induce a significant enhancement of the local electromagnetic(EM) field,which is important for SERS[44,45].As shown in Fig.11,when the dimensions of a metal nanoparticle are less than the wavelength of the incident light,LSPR will occur,leading to collective oscillation of the valence electrons in the metal nanoparticle[46,47].LSPR will concentrate the incident EM field around the nanoparticles.SPPs are the propagating charge oscillations excited by periodicity in a nanostructure.The SPPs will spread along a periodic nanostructure and generate a strong confinement of the EM field[48].However,as SPPs propagate along the surface,they lose energy due to metal absorption,resulting in decay of the EM field[49].Thus,the enhanced Raman signal is the result of LSPR caused by nanoparticles with diameters ranging from 60 to 200 nm generated on the 3D LIPSS,and by SPPs excited by the periodic LIPSS.

Fig.11.Schematic of the SPPs and LSPR caused by laser irradiation during Raman measurement.

4.Conclusions

(1)The microstructure evolution,corrosion behavior,and biocompatibility of a Mg-6Gd-0.6Ca alloy before and after laser surface modification were carefully examined and compared.After laser surface modification,onlyα-Mg remained,and no precipitation of β-phase grains was found in the solidification microstructure.The thickness of the laser-modified layer was estimated to be 403 μm.Dissolution of the β-phase in the laser-melted region resulted in the significant reduction of galvanic couples.In vitro cell culture showed that MC3T3-E1 cells exhibited good adhesion behavior at the surface of all laser-treated specimens;in addition,cells at the laser-melted-and-LIPSS surfaces stretched heavily and presented long filopodia distributed at the outer edge.

赵旭东的《中国山水画中的意义线索与民族志书写》[13]，以中国山水画中的意义线索为例，探讨了中国范式的艺术民族志应该如何书写的问题。这种艺术民族志的书写方式有别于西方人类学科学民族志的主张，是一种通过对景造意表达出来的对意义的深层次阐发，有助于我们重新认识人类学意义的写文化。同时，它为这种写文化的再创造提供了一种可以获得灵感的方法论的新资源，也为中国传统政治语境中权威体系对于自然山水的象征化命名的权力机制提供了一种真正意义上的本土理解。这一理念值得关注。

(2)3D LIPSS decorated with nanoparticles were prepared for SERS on a Ti6Al4V alloy substrate,fabricated by an ultrafast laser in a single step.3D LIPSS provide high enhancement of SERS due to the increased surface area and nanoparticles.After coating with gold film,a significant enhancement of 6×103 was obtained in the Raman measurement.We suggest that the combined effect of LSPR by 3D LIPSS and nanoparticle ‘‘hot spots”is the main reason for the enhancement.

Acknowledgements

This work was supported by the National Key R&D Program of China(2018YFB1107400);the National Key Basic Research Program of China(2015CB059900);the National Natural Science Foundation of China(51705013);and the Beijing Natural Science Foundation(3162019 and J170002).

Compliance with ethics guidelines

Guoqing Hu,Kai Guan,Libin Lu,Jiaru Zhang,Nie Lu,and Yingchun Guan declare that they have no conflict of interest or financial conflicts to disclose.

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