Name: quantitative hardness measurement by instrumented afm-indentation
Uploaded: 2016-11-22T14:00:00.000+00:00
Duration: 8 min 21 s
Description: Watch this Scientific Journal Video about Quantitative Hardness Measurement by Instrumented AFM-indentation at JoVE.com

A.C. is grateful to KoreaTech for financial support.

1. 기악 셋업 및 교정 <ol><li> 경음악 셋업 <ol><li> 0.1 ≥ 180 kHz에서, 품질 계수 Q ≥ (300), 및 굽힘 강성 K ≥ 40 N / m f를 첫 번째 무료 공진 주파수와 유형 DT-NCLR 또는 CDT-NCLR의 단단한 다이아몬드 코팅 캔틸레버를 사용합니다. </li><li> 원자 현미경 제조업체에서 제공 클램프 홀더에 선택한 캔틸레버를 탑재합니다. 긴 축이 AFM의 고속 스캔 방향에 수직이되도록 캔틸레버를 배치 할 특별한주의하십시오. 대안 적으로, 2 성분 에폭시 접착제를 사용하여 AFM 제조업체에서 제공 캔틸레버 홀더에 캔틸레버 접착제. </li><li> 원자 현미경 헤드에 캔틸레버 홀더를 장착하고 AFM 캔틸레버에 초점을 AFM 시스템에 일반적으로 사용할 수있는 광학 현미경을 사용합니다. 캔틸레버의 긴 축이 고속 스캔 방향에 수직임을 다시 한 번 확인. 그렇지 않은 경우, 뒤로 이동제 1.1.2. </li><li> 이 캔틸레버의 끝 부분에서 반사되도록 레이저 빔을 맞추고. 포토 다이오드에서의 전압의 합을 모니터 및 합 신호를 최대화하도록 미세 조정을 수행. 전형적인 합 신호 값이 V. 범위에 </li><li> 수직 및 횡 방향의 변위에 대응하는 전압이 거의 제로가되어, 포토 다이오드의 중심으로 반사 된 레이저 스폿을 가지고하도록 미러의 수평 및 수직 방향의 경사각을 조정한다. </li></ol></li><li> 구경 측정 <ol><li> 캔틸레버 0,1 f를 제 자유 굽힘 공진을 결정하기 위해 주파수 스위프를 수행한다. </li><li> 19에 따라 계산 캔틸레버 (K)의 휨 강성을 결정 (1) <img alt="식 (1)" src="/files/ftp_upload/54706/54706eq1.jpg" /> E는 영률이며 여기서 cantil의 폭 w는, L은, 캔틸레버의 길이지금까지, T는 두께이다. 이를 위해, 더 나은 정확성을 광학 현미경 또는 주사 전자 현미경에 의한 캔틸레버의 길이 및 폭을 측정한다. 항에있어서, f를 0.1 최초 자유 굽힘 공진 주파수에서 캔틸레버의 두께를 계산 (2) <img alt="식 (2)" src="/files/ftp_upload/54706/54706eq2.jpg" /> 여기서, ρ는 질량 밀도이다. </li><li> 특정 캔틸레버 타입의 포토 다이오드 감도의 기본값을 선택 강자성의 설정 메뉴의 실험에 사용한다. 접근 버튼을 클릭하여 10 윈를로드 F에서 기준 샘플과의 접촉에 캔틸레버 팁을 가져 N =. </li><li> 원자 현미경 소프트웨어의 힘 분광 메뉴를 열고 50 nm의 0.3 μm의 / 초에 Z-스캐너 수축 / 확장에 Z-스캐너의 상대적 수축과 확장을 설정합니다. 그래서 힘 - 거리 곡선의 기록을 수행 할멀리 시료 표면에서 다음 같은 거리의 접근 및 철회의 시리즈의 50 나노 미터에 Z-스캐너의 철회의 첫 번째 구성됩니다. </li><li> 샘플 변형 효과를 피하기 위하여, 나노 결정질 다이아몬드 또는 사파이어와 같은 부드럽고 부적합한면 1.2.4 제안 설정된 파라미터와 힘 거리 커브를 기록한다. 이렇게하려면 때문에 AFM 소프트웨어의 힘 분광 메뉴에서 획득 버튼을 클릭합니다. </li><li> 강자성 소프트웨어 보정 메뉴에서 선형 함수로 힘 - 거리 곡선의 반발력 부를 장착한다. 피팅 라인의 역 기울기는 포토 다이오드 감도 S에 대응한다. 교정 실행] 버튼을 클릭하여 AFM 소프트웨어 보정 메뉴의 장비 소프트웨어의 기본 값으로 결정된 값을 대체. </li></ol></li></ol> 2. 샘플 준비 참고 : 생 측정 샘플S 실험 물리적 기상 증착에 의해 운모 성장을 100 nm의 두께의 Au 아주 매끄러운 (111)의 박막 구성된다. <ol><li> 양면 카본 테이프를 이용하여 기기의 제조자에 의해 제공된 자기 시료 홀더 상에 샘플을 탑재. 카본 테이프 긴장하게하도록 측정시 샘플의 드리프트를 피하기 위해, 측정 전에 샘플을 하루 마운트. 또한, 일반적으로 몇 분 이내에 건조 실버 페인트와 홀더에 샘플을 장착합니다. </li><li> 은 x / y를 스캐너에 자기 샘플 홀더를 탑재합니다. </li></ol> 3. 측정 절차 <ol><li> 오프 공명이 값이 자동으로이 특정 캔틸레버의 악기 소프트웨어에 의해 설정되어 있는지 A = 20 nm의 주에서 발진 진폭 (이 실험 F = 190.67 kHz에서에서) 약간 발진 주파수를 설정합니다. 세트 포인트 = 5 nm에서 수동으로 발진 세트 포인트를 설정합니다. </li><li> 무승부원자 현미경의 스텝 모터를 사용하여 샘플의 표면을 향해 캔틸레버. 힘 센서는 시료 표면과 충돌하지 않는지 확인합니다. 거친 방법 중 초점 캔틸레버를 유지하고 시료 표면이 완벽하게 초점이 전에 거친 접근 방식을 중지합니다. </li><li> 자동 방식 버튼을 클릭하여 힘 센서 접근. 진동 진폭이 설정치에 도달하면, 선단이 시료 표면의 지형을 스캔 할 준비가되어있다. </li><li> 5 × 5 내지 1 × 1 μm²에 이르는 영역에서 지형 이미지들의 시리즈를 녹화 (가능한 경우, X / Y 스캐너를 기울 지형 신호의 기울기를 조정). 같은 지역의 연속적인 이미지 드리프트의 흔적을 전시하지 않는 것이 및 Z 스캐너의 위치가 거의 일정하게 유지해야합니다. 그렇지 않은 경우, 시스템이 안정화 될 때까지 촬영을 계속한다. </li><li> 시스템이 안정하고 평활 한 1 × 1 μm² 영역이 발견되면, F 후퇴ORCE는 후퇴 버튼을 클릭하여 시료 표면에서 몇 마이크로 미터 센서. </li><li> 악기 메뉴의 힘 분광 모드를 선택하고 10 nm의 힘 설정 점과 함께 미리 선택된 1 × 1 μm² 지역의 중간에 힘 센서를 이동합니다. 이 일정하게 유지 될 때까지 Z 스캐너의 위치를 ​​모니터한다. </li><li> 중심 미리 선택된 1 × 1 μm² 지역의 중심에 해당하는 포인트의 2 × 2 그리드를 선택합니다. 500 nm에서이 옆에 이웃 점 사이의 거리를 설정합니다. </li><li> 300 ㎚ / sec의 속도로 0 내지 150nm에서 변화시키는 스캐너 상대적인 거리를 설정하고 동일 거리에 걸쳐 동일한 속도로 후퇴. 시료 표면에 대하여 캔틸레버의 경사각을 주어, φ는 경사각 20 수직 스캐너 연장 Z 동안 황갈색 φ × Z 의한 횡 스캐너를 이동하여 틸트 보정을 적용한다. 참고 : 몇 이달ruments 자신의 힘 분광법 또는 들여 쓰기 모드에서 캔틸레버 기울기를 차지; 이 작품에 사용 된 AFM의 경우입니다. </li><li> 원자 현미경 들여 쓰기 데이터의 수집을 시작하는 계기 소프트웨어의 시작 버튼을 누릅니다. </li><li> 강자성 압입 측정이 완료되면, 마이크로 미터 떨어진 샘플 표면으로부터의 힘 센서를 철회. </li><li> 계기 소프트웨어 메뉴 비접촉 AFM 모드 영상을 선택하고 섹션 3.1 및 3.2에 기술 된 절차를 반복한다. </li><li> 그래서 들여 쓰기의 정확한 위치를 찾는으로 3.3 절에서와 동일한 1 × 1 μm² 표면 영역에 걸쳐 검사를 수행합니다. 500 X 500 nm² 표면적 위에 또한 표면 검사보다 상세한 이미지를 나머지 압흔을 행할 수있다. </li></ol> 4. 데이터 분석 <ol><li> 이미지 처리 <ol><li> 고속 스캔 DIR의 라인을 정렬하도록 기록 지형 이미지를 처리평균 차이에 기초 ection. Gwyddion의 내장 함수를 사용합니다. </li></ol></li><li> 투영 면적을 Gwyddion의 들여 쓰기 분석 기능을 사용하여 들여 쓰기의 페이지를 계산합니다. </li><li> Gwyddion의 선단 분석 기능을 사용하여 압흔의 지형 화상으로부터 AFM 팁 형상을 추정한다. 그런 다음 팁 형상 이미지를 평균하여 평균 팁 형상의 반 개구 각 α를 측정한다. </li><li> 팁 변위 δ를 산출하여 (13)에 따른 힘 - 변위 곡선에 힘 - 거리 곡선 변환 (삼) <img alt="식 (3)" src="/files/ftp_upload/54706/54706eq3.jpg" /> 여기서 Z는 스캐너 상대적인 위치이다. </li><li> 이제 팁 변위 대 힘을 플롯. 그 결과 곡선은 일반적으로 원자의 소성 이벤트에 해당하는 몇 백 오후 범위의 길이로, 소위 팝업 기능을 표시합니다. 살전의 첫 번째를 사용전자 팝업 기능은 엘 4 δ 탄성 한계에 팁 변위를 확인합니다. </li><li> 헤르츠 기능 (21)와 힘 - 변위 곡선의 탄성 부분을 맞 춥니 다. (4) <img alt="식 (4)" src="/files/ftp_upload/54706/54706eq4.jpg" /> 어디 R은 팁 반경 및 E &#39;*에 의해 주어진, 탄력의 감소 계수이다 <img alt="식 (5)" src="/files/ftp_upload/54706/54706eq5.jpg" /> , M은 S, t는 각각의 샘플과 팁의 압입 탄성률 되 고. 이 경우, 적합 파라미터이다 <img alt="식 (6)" src="/files/ftp_upload/54706/54706eq6.jpg" /> . </li><li> 피팅 함수 및 실험 곡선 (21) 사이의 면적 차이로부터 소성 W 소성의 작업을 계산하도록 가소성 체제에 맞는 기능을 확장한다. </li><li> 1에 따른 2 샘플의 경도를 계산 (5) <img alt="식 (7)" src="/files/ftp_upload/54706/54706eq7.jpg" /> 과 (6) <img alt="식 (8)" src="/files/ftp_upload/54706/54706eq8.jpg" /> F의 N이 최대가 최대 하중이고,는 P 섹션 4.2에서 계산 된 덴트의 투영 면적이, α는 4.3에서 계산 된 팁의 반 개방 각도이고, δ EL은 제 가소성의 팁 변위이고 이벤트 및 δ 최대는 최대 팁 변위 (4.4 절 참조)이다. </li></ol>

<ol>
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M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=On+the+use+of+nanoscale+indentation+with+the+AFM+in+the+identification+of+phases+in+blends+of+linear+low+density+polyethylene+and+high+density+polyethylene.">On the use of nanoscale indentation with the AFM in the identification of phases in blends of linear low density polyethylene and high density polyethylene.</a> J. Mater. Sci. 35 (1), 221-228 (2000).</li><li>Zhang, L., Wang, W., Zheng, L., Wang, X., Yan, Q. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantitative+characterization+of+mechanical+property+of+annealed+monolayer+colloidal+crystal.">Quantitative characterization of mechanical property of annealed monolayer colloidal crystal.</a> Langmuir. 32 (2), 451-459 (2016).</li><li>Ne&#269;as, D., Klapetek, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gwyddion:+An+open-source+software+for+SPM+data+analysis.">Gwyddion: An open-source software for SPM data analysis.</a> Cent. Eur. J. Phys. 10 (1), 181-188 (2012).</li><li>Hahn, B. H., Valentine, D. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Essential Matlab for Engineers and Scientists. , (2013).</li><li>Nonnenmacher, M., Greschner, J., Wolter, O., Kassing, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scanning+Force+Microscopy+with+Micromachined+Silicon+Sensors.">Scanning Force Microscopy with Micromachined Silicon Sensors.</a> J. Vac. Sci. Technol. B. 9 (2), 1358-1362 (1991).</li><li>Cannara, R. J., Brukman, M. J., Carpick, R. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cantilever+tilt+compensation+for+variable-load+atomic+force+microscopy.">Cantilever tilt compensation for variable-load atomic force microscopy.</a> Rev. Sci. Instrum. 76 (5), 053706 (2005).</li><li>Johnson, K. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Contact Mechanics. , (1985).</li><li>Mohr, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Young's+Modulus,+Fracture+Strength,+and+Poisson's+Ratio+of+Nanocrystalline+Diamond+Films.">Young's Modulus, Fracture Strength, and Poisson's Ratio of Nanocrystalline Diamond Films.</a> J. Appl. Phys. 116 (12), 124308-1-124308-10 (2014).</li><li>Arnault, J. C., Mosser, A., Zamfirescu, M., Pelletier, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Elastic+recovery+measurements+performed+by+atomic+force+microscopy+and+standard+nanoindentation+on+a+Co(10.1)+monocrystal.">Elastic recovery measurements performed by atomic force microscopy and standard nanoindentation on a Co(10.1) monocrystal.</a> J. Mater. Res. 17 (6), 1258-1265 (2002).</li><li>Cao, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nanoindentation+measurements+of+the+mechanical+properties+of+polycrystalline+Au+and+Ag+thin+films+on+silicon+substrates:+Effect+of+grain+size+and+film+thickness.">Nanoindentation measurements of the mechanical properties of polycrystalline Au and Ag thin films on silicon substrates: Effect of grain size and film thickness.</a> Mater. Sci. Eng. A. 457 (1-2), 232-240 (2006).</li><li>Lilleodden, E. T., Nix, W. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microstructural+length-scale+effects+in+the+nanoindentation+behavior+of+thin+gold+films.">Microstructural length-scale effects in the nanoindentation behavior of thin gold films.</a> Acta Mater. 54 (6), 1583-1593 (2006).</li><li>Corcoran, S. G., Colton, R. J., Lilleodden, E. T., Gerberich, W. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anomalous+plastic+deformation+at+surfaces:+Nanoindentation+of+gold+single+crystals.">Anomalous plastic deformation at surfaces: Nanoindentation of gold single crystals.</a> Phys. Rev. B. 55 (24), R16057 (1997).</li><li>Van Vliet, K. J., Li, J., Zhu, T., Yip, S., Suresh, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantifying+the+early+stages+of+plasticity+through+nanoscale+experiments+and+simulations.">Quantifying the early stages of plasticity through nanoscale experiments and simulations.</a> Phy. Rev. B. 67 (10), 104105 (2003).</li></ol>

The authors have nothing to disclose.

방법은 다이아몬드 코팅 된 AFM 팁을 갖는 금 (111)에 박막의 표면 요철의 시리즈를 수행하기 위해 제공되었다. 비접촉 AFM 이미징 AFM 오목 동일한 하중 센서로 실시 하였다. 비접촉 촬상 대한 요구가 0.1 ≥ 180 kHz 및 AFM 들여에서 높은 품질 계수 Q ≥ 300 F 높은 제 자유 공진 주파수이고, 수직 힘이 몇 마이크로 뉴턴의 범위에 적용되기 위해서는 높은 굽힘 강성 캔틸레버가 ?...

For the last 150 years, hardness values have been used to characterize the mechanical behavior of materials and to predict their performance under various loading conditions. The hardness of a material describes the resistance of its surface to the penetration of a harder indenter. For metallic materials, the hardness relates to resistance to plastic flow.
Although the implementation of hardness testing has proven to be relatively easy, the yielded results have long been rather empirical, making them unable to describe the intrinsic properties of the investigated material. The empirical nature of hardness testing results has been a consequence of the use of various indenter geometries, each of which has a different established hardness scale. However, all empirical hardness scales, such as the Brinell hardness (HB) test for spherical indenters and the Vickers (HV) and Knoop (HK) hardness tests for different types of four-sided pyramids, have one thing in common: the hardness number is determined from the ratio of the applied load to the developed area of the remaining indent. In an attempt to relate the hardness of metals to their fundamental mechanical properties, the hardness measured with a spherical indenter has been defined as the ratio of the applied load to the projected area of the remaining indent. This hardness value, also known as the Meyer's hardness (H), is equal to the mean contact pressure and directly relates to the yield strength of non-work-hardening metals1. 
Hardness testing has long been limited to the macro-scale, in which case the sizes of indents have been measured by optical means. The development of instrumented indentation, where force-displacement curves are recorded, has been recognized as a valuable alternative for determining the hardness of a material, although the size of the remaining indent may be too small to be accurately imaged. In this case, the projected area of the indent is calculated from the tip displacement according to a so-called indenter-area function2. Due to this development, the analysis of the curvature of force-displacement curves and of the occurrence of distinct plasticity events in the shape of pop-ins is now widely used to study the mechanisms of plastic deformation at the micrometer scale, such as by means of nanoindentation3.
It is well accepted that the carriers of plastic deformation in metals, i.e., dislocations, operate at the nanometer scale. To understand their modes of operation at the atomic level, new experimental techniques with atomic resolution are needed. Initial investigations of atomistic plasticity events at the interfaces between nanometer-sized single asperities and single crystalline Au surfaces of different orientations have been carried out by interfacial force microscopy (IFM)4, 5. Atomic force microscopy (AFM) indentation has been applied to observe the nucleation and gliding of single dislocations in KBr(100) single crystals6, Cu(100)7, and Au(111)8, 9. There, atomistic plasticity events were observed in the shape of pop-ins, with lengths in the range of 1 &#197;. AFM indentation has also been used to measure the nano-hardness and -elasticity of gold nano-island grown on mica10. In this work, the nano-hardness was found to be smaller than the bulk value and was observed to depend linearly on the indentation area. Also, a detailed measurement protocol has been proposed to determine the hardness of hard surfaces, such as fused quartz and silicon, by AFM indentation with a diamond-tipped sapphire cantilever11. In particular, this method takes into account the non-linearity of the photo-sensitive deflection detectors for large cantilever deflection12. More recently, AFM indentation and non-contact AFM imaging have been combined to quantitatively determine the hardness and the fundamental mechanisms of plastic deformation of a Pt(111) single crystalline surface and Pt-based metallic glass13. For Pt(111), plastic deformation mechanisms at the nanometer scale were found to be consistent with the discrete mechanisms established for larger scales. Further, the nanometer-scale plastic deformation of the metallic glass was found to be not discrete, but rather continuous and highly localized around the indenting tip. These results revealed a lower size limit for metallic glasses, below which shear transformation mechanisms are not activated by indentation. AFM indentation has also been used to determine the hardness values of biological samples (such as collagen fibrils)14, polymers15, and colloidal crystals16.
In the present work, a careful experimental procedure is described that includes the force sensor selection, its calibration, the calibration of the cantilever deflection detection system, and the minimization of instrumental drift for accurate and reproducible force-distance measurements. Also, a method for the data analysis is presented that allows the extraction of force-penetration curves from recorded force-distance curves. Further representative results are shown and discussed in the light of recent findings in the field of plastic deformation of small volumes.
For this experiment, an atomic force microscope (AFM) was used. The cornerstone of an AFM is its micro-fabricated force sensor (usually a cantilever beam), with a sharp tip at the end whose radius is in the range of several nanometers. In the particular configuration of the instrument used for this experiment, the cantilever is mounted onto a piezo-electric z-scanner, while the sample to be investigated is mounted onto a piezo-electric x/y-scanner. During AFM imaging, the force sensor tip is scanned over a sample to register interaction forces with the sample surface that result in a deflection of the cantilever according to Hooke's law. The static or dynamic deflection of the cantilever can be measured with an optical beam deflection detector, consisting of a laser diode, a set of mirrors, and a photodiode that converts the cantilever deflection into a voltage. During imaging, the signal at the photodiode is controlled by a feedback loop so as to keep the interaction forces at the sample surface; this results in adjustments of the z-scanner position, which are recorded and displayed as a topography image.
Prerequisites for the experiment described below are that the piezo-electric scanners of the atomic force microscope are well-calibrated and that the instrument stays in the laboratory at a constant temperature and humidity level. The reader should be aware that, depending on the atomic force microscope model, some of the experimental procedure steps may have to be modified. In particular, all measurements are performed after setting the scanners' ranges to "small"; this function allows for the reduction of the x/y-scanner range to 5 x 5 &#181;m&#178; and the z-scanner range to 4 &#181;m, which ensures a resolution at small scales. This function is not available on all commercial AFM and is not mentioned in the remainder of the text.
For data analysis, the use of the free SPM data analysis software Gwyddion17 and the Matlab software package18 are recommended.
This protocol gives a description of the experimental procedure to be followed in order to perform instrumented hardness measurements by AFM. Since the handling of different commercial AFMs may differ from one model to the other, the reader should refer to the manual provided by the AFM manufacturer for detailed setting procedures and software information. In the following text, in order to reproduce the described experiment, it is assumed that the reader is familiar with the handling of the particular AFM used here.

<table><tbody><tr><td>AFM XE-100</td><td>Park Instruments</td><td>discontinued</td><td>Atomic force microscope</td></tr><tr><td>CDT-NCLR</td><td>NanoSensors</td><td>CDT-NCLR</td><td>Conductive diamond coated non-contact lever</td></tr><tr><td>100 nm thick Au(111) thin film on Mica</td><td>Phasis</td><td>20020011</td><td>atomically smooth gold thin film</td></tr></tbody></table>

quantitative hardness measurement by instrumented afm-indentation

In this work, a combination of amplitude-modulated non-contact atomic force microscopy and atomic force spectroscopy is applied for instrumented hardness measurements on an Au(111) surface with atomistic resolution of single plasticity events. A careful experimental procedure is described that includes the force sensor selection, its calibration, the calibration of the cantilever deflection detection system, and the minimization of instrumental drift for accurate and reproducible force-distance measurements. Also, a method for the data analysis is presented that allows the extraction of force-penetration curves from recorded force-distance curves. A typical curve displays a clear elastic deformation regime up to the first plasticity event, or pop-in, with a length in the range of one to two Burger's vectors. Later plasticity events exhibit the same magnitude. The work of plasticity is further extracted from the measurements. Finally, the hardness is determined in combination with the indentation curve using non-contact atomic force microscopy images of the remaining indents.

본 연구에서는 캔틸레버 (K)의 굽힘 강성이 기하학적 빔 이론 (19)에 따라 계산 하였다. 이 연구에서 사용 된 특정 다이아몬드 코팅 캔틸레버를 들어, 우리는 K = 55.69 N / m를 발견했다. 우리는 다이아몬드 코팅을 무시합니다; 다이아몬드 막의 두께는 (그 영률은 실리콘보다 훨씬 더 크다하더라도) 대폭 굽힘 강성을 증가시키지 않고 따라서 캔틸레버 두께...

Watch this Scientific Journal Video about Quantitative Hardness Measurement by Instrumented AFM-indentation at JoVE.com

Quantitative Hardness Measurement by Instrumented AFM-indentation

This experimental protocol describes how atomic force microscopy can be used to measure hardness at the true nanometer scale and to detect single atomistic plasticity events.

계장 AFM-들여 쓰기에 의해 정량 경도 측정

In this work, a combination of amplitude-modulated non-contact atomic force microscopy and atomic force spectroscopy is applied for instrumented hardness ...

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9.6K 조회수.  KoreaTech, Korea University of Technology and Education. This experimental protocol describes how atomic force microscopy can be used to measure hardness at the true nanometer scale and to detect single atomistic plasticity events. 

비디오: Quantitative Hardness Measurement by Instrumented AFM-indentation

Measuring the Mechanical Properties of Living Cells Using Atomic Force Microscopy

Mechanical properties of cells and extracellular matrix (ECM) play important roles in many biological processes including stem cell differentiation, tumor formation, and wound healing. Changes in stiffness of cells and ECM are often signs of changes in cell physiology or diseases in tissues. Hence, cell stiffness is an index to evaluate the status of cell cultures. Among the multitude of methods applied to measure the stiffness of cells and tissues, micro-indentation using an Atomic Force Microscope (AFM) provides a way to reliably measure the stiffness of living cells. This method has been widely applied to characterize the micro-scale stiffness for a variety of materials ranging from metal surfaces to soft biological tissues and cells. The basic principle of this method is to indent a cell with an AFM tip of selected geometry and measure the applied force from the bending of the AFM cantilever. Fitting the force-indentation curve to the Hertz model for the corresponding tip geometry can give quantitative measurements of material stiffness. This paper demonstrates the procedure to characterize the stiffness of living cells using AFM. Key steps including the process of AFM calibration, force-curve acquisition, and data analysis using a MATLAB routine are demonstrated. Limitations of this method are also discussed.

This paper demonstrates a protocol to characterize the mechanical properties of living cells by means of microindentation using an Atomic Force Microscope (AFM).

원자 힘 현미경을 사용하여 살아있는 세포의 기계적 특성을 측정

Mechanical  properties of cells and extracellular matrix (ECM) play important roles in many  biological processes including stem cell differentiation, ...

연구

생체공학

Characterization Of Multi-layered Fish Scales (Atractosteus spatula) Using Nanoindentation, X-ray CT, FTIR, and SEM

The hierarchical architecture of protective biological materials such as mineralized fish scales, gastropod shells, ram&#8217;s horn, antlers, and turtle shells provides unique design principles with potentials for guiding the design of protective materials and systems in the future. Understanding the structure-property relationships for these material systems at the microscale and nanoscale where failure initiates is essential. Currently, experimental techniques such as nanoindentation, X-ray CT, and SEM provide researchers with a way to correlate the mechanical behavior with hierarchical microstructures of these material systems1-6. However, a well-defined standard procedure for specimen preparation of mineralized biomaterials is not currently available. In this study, the methods for probing spatially correlated chemical, structural, and mechanical properties of the multilayered scale of A. spatula using nanoindentation, FTIR, SEM, with energy-dispersive X-ray (EDX) microanalysis, and X-ray CT are presented.

Characterization Of Multi-layered Fish Scales Atractosteus spatula Using Nanoindentation, X-ray CT, FTIR, and SEM

This paper presents the methods used for probing spatially correlated chemical, structural, and mechanical properties of the multilayered scale of Atractosteus spatula (A. spatula) using nanoindentation, Fourier transform infrared (FTIR) spectroscopy, scanning electron microscopy (SEM), and X-ray computed tomography (X-ray CT). The experimental results have been used to investigate the design principles of protective biological materials.

다층 물고기 비늘 (의 특성<em&gt; Atractosteus 주걱</em&gt;) 나노 인 덴 테이션, X-선 CT, FTIR, 및 SEM을 사용하여

The hierarchical architecture of protective biological materials such as mineralized fish scales, gastropod shells, ram&#8217;s horn, antlers, and turtle ...

Use of Atomic Force Microscopy to Measure Mechanical Properties and Turgor Pressure of Plant Cells and Plant Tissues

We present here the use of atomic force microscopy to indent plant tissues and recover its mechanical properties. Using two different microscopes in indentation mode, we show how to measure an elastic modulus and use it to evaluate cell wall mechanical properties. In addition, we also explain how to evaluate turgor pressure. The main advantages of atomic force microscopy are that it is non-invasive, relatively rapid (5~20 min), and that virtually any type of living plant tissue that is superficially flat can be analyzed without the need for treatment. The resolution can be very good, depending on the tip size and on the number of measurements per unit area. One limitation of this method is that it only gives direct access to the superficial cell layer.

Here, we present atomic force microscopy (AFM), operated as a nano- and micro-indentation tool on cells and tissues. The instrument allows the simultaneous acquisition of 3D surface topography of the sample and its mechanical properties, including cell wall Young's modulus as well as turgor pressure.

식물 세포 및 식물 조직의 기계적 특성 및 터거 압력을 측정하기 위해 원자력 현미경 검사법 사용

We present here the use of atomic force microscopy to indent plant tissues and recover its mechanical properties. Using two different microscopes in ...

발생학

Covalent Attachment of Single Molecules for AFM-based Force Spectroscopy

Atomic force microscopy (AFM)-based single molecule force spectroscopy is an ideal tool for investigating the interactions between a single polymer and surfaces. For a true single molecule experiment, covalent attachment of the probe molecule is essential because only then can hundreds of force-extension traces with one and the same single molecule be obtained. Many traces are in turn necessary to prove that a single molecule alone is probed. Additionally, passivation is crucial for preventing unwanted interactions between the single probe molecule and the AFM cantilever tip as well as between the AFM cantilever tip and the underlying surface. The functionalization protocol presented here is reliable and can easily be applied to a variety of polymers. Characteristic single molecule events (i.e., stretches and plateaus) are detected in the force-extension traces. From these events, physical parameters such as stretching force, desorption force and desorption length can be obtained. This is particularly important for the precise investigation of stimuli-responsive systems at the single molecule level. As exemplary systems poly(ethylene glycol) (PEG), poly(N-isopropylacrylamide) (PNiPAM) and polystyrene (PS) are stretched and desorbed from SiOx (for PEG and PNiPAM) and from hydrophobic self-assembled monolayer surfaces (for PS) in aqueous environment.

Covalent attachment of probe molecules to atomic force microscopy (AFM) cantilever tips is an essential technique for the investigation of their physical properties. This allows us to determine the stretching force, desorption force and length of polymers via AFM-based single molecule force spectroscopy with high reproducibility.

AFM 기반 힘 분광학을 위한 단일 분자의 공유 부착

Atomic force microscopy (AFM)-based single molecule force spectroscopy is an ideal tool for investigating the interactions between a single polymer and ...

화학

Atomic Force Microscopy Cantilever-Based Nanoindentation: Mechanical Property Measurements at the Nanoscale in Air and Fluid

An atomic force microscope (AFM) fundamentally measures the interaction between a nanoscale AFM probe tip and the sample surface. If the force applied by the probe tip and its contact area with the sample can be quantified, it is possible to determine the nanoscale mechanical properties (e.g., elastic or Young&#39;s modulus) of the surface being probed. A detailed procedure for performing quantitative AFM cantilever-based nanoindentation experiments is provided here, with representative examples of how the technique can be applied to determine the elastic moduli of a wide variety of sample types, ranging from kPa to GPa. These include live mesenchymal stem cells (MSCs) and nuclei in physiological buffer, resin-embedded dehydrated loblolly pine cross-sections, and Bakken shales of varying composition.
Additionally, AFM cantilever-based nanoindentation is used to probe the rupture strength (i.e., breakthrough force) of phospholipid bilayers. Important practical considerations such as method choice and development, probe selection and calibration, region of interest identification, sample heterogeneity, feature size and aspect ratio, tip wear, surface roughness, and data analysis and measurement statistics are discussed to aid proper implementation of the technique. Finally, co-localization of AFM-derived nanomechanical maps with electron microscopy techniques that provide additional information regarding elemental composition is demonstrated.

Quantifying the contact area and force applied by an atomic force microscope (AFM) probe tip to a sample surface enables nanoscale mechanical property determination. Best practices to implement AFM cantilever-based nanoindentation in air or fluid on soft and hard samples to measure elastic modulus or other nanomechanical properties are discussed.

An atomic force microscope (AFM) fundamentally measures the interaction between a nanoscale AFM probe tip and the sample surface. If the force applied by ...

공학

Characterizing Multiscale Mechanical Properties of Brain Tissue Using Atomic Force Microscopy, Impact Indentation, and Rheometry

To design and engineer materials inspired by the properties of the brain, whether for mechanical simulants or for tissue regeneration studies, the brain tissue itself must be well characterized at various length and time scales. Like many biological tissues, brain tissue exhibits a complex, hierarchical structure. However, in contrast to most other tissues, brain is of very low mechanical stiffness, with Young&#39;s elastic moduli E on the order of 100s of Pa. This low stiffness can present challenges to experimental characterization of key mechanical properties. Here, we demonstrate several mechanical characterization techniques that have been adapted to measure the elastic and viscoelastic properties of hydrated, compliant biological materials such as brain tissue, at different length scales and loading rates. At the microscale, we conduct creep-compliance and force relaxation experiments using atomic force microscope-enabled indentation. At the mesoscale, we perform impact indentation experiments using a pendulum-based instrumented indenter. At the macroscale, we conduct parallel plate rheometry to quantify the frequency dependent shear elastic moduli. We also discuss the challenges and limitations associated with each method. Together these techniques enable an in-depth mechanical characterization of brain tissue that can be used to better understand the structure of brain and to engineer bio-inspired materials.

We present a set of techniques to characterize the viscoelastic mechanical properties of brain at the micro-, meso-, and macro-scales.

원자 힘 현미경, 충격 들여 쓰기 및 유변학를 사용하여 뇌 조직의 멀티 스케일 기계 속성을 특성화

To design and engineer materials inspired by the properties of the brain, whether for mechanical simulants or for tissue regeneration studies, the brain ...

신경과학

Micro-Mechanical Characterization of Lung Tissue Using Atomic Force Microscopy

Matrix stiffness strongly influences growth, differentiation and function of adherent cells1-3. On the macro scale the stiffness of tissues and organs within the human body span several orders of magnitude4. Much less is known about how stiffness varies spatially within tissues, and what the scope and spatial scale of stiffness changes are in disease processes that result in tissue remodeling. To better understand how changes in matrix stiffness contribute to cellular physiology in health and disease, measurements of tissue stiffness obtained at a spatial scale relevant to resident cells are needed. This is particularly true for the lung, a highly compliant and elastic tissue in which matrix remodeling is a prominent feature in diseases such as asthma, emphysema, hypertension and fibrosis. To characterize the local mechanical environment of lung parenchyma at a spatial scale relevant to resident cells, we have developed methods to directly measure the local elastic properties of fresh murine lung tissue using atomic force microscopy (AFM) microindentation. With appropriate choice of AFM indentor, cantilever, and indentation depth, these methods allow measurements of local tissue shear modulus in parallel with phase contrast and fluorescence imaging of the region of interest. Systematic sampling of tissue strips provides maps of tissue mechanical properties that reveal local spatial variations in shear modulus. Correlations between mechanical properties and underlying anatomical and pathological features illustrate how stiffness varies with matrix deposition in fibrosis. These methods can be extended to other soft tissues and disease processes to reveal how local tissue mechanical properties vary across space and disease progression.

The stiffness of the extracellular matrix strongly influences multiple behaviors of adherent cells. Matrix stiffness varies spatially throughout a tissue, and undergoes modification in various disease conditions. Here we develop methods to characterize spatial variations in stiffness in normal and fibrotic mouse lung tissue using atomic force microscopy microindentation.

원자 힘 현미경을 사용하여 폐 조직의 미세 기계적 특성

Matrix stiffness strongly influences growth, differentiation and function of adherent cells1-3.  On the macro scale the stiffness of tissues and organs ...

생물학

Application of Atomic Force Microscopy to Detect Early Osteoarthritis

Biomechanical properties of cells and tissues not only regulate their shape and function but are also crucial for maintaining their vitality. Changes in elasticity can propagate or trigger the onset of major diseases like cancer or osteoarthritis (OA). Atomic force microscopy (AFM) has emerged as a strong tool to qualitatively and quantitatively characterize the biomechanical properties of specific biological target structures on a microscopic scale, measuring forces in a range from as small as the piconewton to the micronewton. Biomechanical properties are of special importance in musculoskeletal tissues, which are subjected to high levels of strain. OA as a degenerative disease of the cartilage results in the disruption of the pericellular matrix (PCM) and the spatial rearrangement of the chondrocytes embedded in their extracellular matrix (ECM). Disruption in PCM and ECM has been associated with changes in the biomechanical properties of cartilage. In the present study we used AFM to quantify these changes in relation to the specific spatial pattern changes of the chondrocytes. With each pattern change, significant changes in elasticity were observed for both the PCM and ECM. Measuring the local elasticity thus allows for drawing direct conclusions about the degree of local tissue degeneration in OA.

We present a method to investigate early osteoarthritic changes at the cellular level in articular cartilage by using atomic force microscopy (AFM).

조기 골관절염을 검출하기 위한 원자력 현미경 검사법 적용

Biomechanical properties of cells and tissues not only regulate their shape and function but are also crucial for maintaining their vitality. Changes in ...

Experimental and Data Analysis Workflow for Soft Matter Nanoindentation

Nanoindentation refers to a class of experimental techniques where a micrometric force probe is used to quantify the local mechanical properties of soft biomaterials and cells. This approach has gained a central role in the fields of mechanobiology, biomaterials design&#160;and tissue engineering, to obtain a proper mechanical characterization of soft materials with a resolution comparable to the size of single cells (&#956;m). The most popular strategy to acquire such experimental data is to employ an atomic force microscope (AFM); while this instrument offers an unprecedented resolution in force (down to pN) and space (sub-nm), its usability is often limited by its complexity that prevents routine measurements of integral indicators of mechanical properties, such as Young&#39;s Modulus (E). A new generation of nanoindenters, such as those based on optical fiber sensing technology, has recently gained popularity for its ease of integration while allowing to apply sub-nN forces with &#181;m spatial resolution, therefore being suitable to probe local mechanical properties of hydrogels and cells.
In this protocol, a step-by-step guide detailing the experimental procedure to acquire nanoindentation data on hydrogels and cells using a commercially available ferrule-top optical fiber sensing nanoindenter is presented. Whereas some steps are specific to the instrument used herein, the proposed protocol can be taken as a guide for other nanoindentation devices, granted some steps are adapted according to the manufacturer&#39;s guidelines. Further, a new open-source Python software equipped with a user-friendly graphical user interface for the analysis of nanoindentation data is presented, which allows for screening of incorrectly acquired curves, data filtering, computation of the contact point through different numerical procedures, the conventional computation of E, as well as a more advanced analysis particularly suited for single-cell nanoindentation data.

The protocol presents a complete workflow for soft material nanoindentation experiments, including hydrogels and cells. First, the experimental steps to acquire force spectroscopy data are detailed; then, the analysis of such data is detailed through a newly developed open-source Python software, which is free to download from GitHub.

연질 물질 나노인덴테이션을 위한 실험 및 데이터 분석 워크플로우

Nanoindentation refers to a class of experimental techniques where a micrometric force probe is used to quantify the local mechanical properties of soft ...

Addressing Practical Issues in Atomic Force Microscopy-Based Micro-Indentation on Human Articular Cartilage Explants

Without a doubt, atomic force microscopy (AFM) is currently one of the most powerful and useful techniques to assess micro and even nano-cues in the biological field. However, as with any other microscopic approach, methodological challenges can arise. In particular, the characteristics of the sample, sample preparation, type of instrument, and indentation probe can lead to unwanted artifacts. In this protocol, we exemplify these emerging issues on healthy as well as osteoarthritic articular cartilage explants. To this end, we first show via a step-by-step approach how to generate, grade, and visually classify ex vivo articular cartilage discs according to different stages of degeneration by means of large 2D mosaic fluorescence imaging of the whole tissue explants. The major strength of the ex vivo model is that it comprises aged, native, human cartilage that allows the investigation of osteoarthritis-related changes from early onset to progression. In addition, common pitfalls in tissue preparation, as well as the actual AFM procedure together with the subsequent data analysis, are also presented. We show how basic but crucial steps such as sample preparation and processing, topographic sample characteristics caused by advanced degeneration, and sample-tip interaction can impact data acquisition. We also subject to scrutiny the most common problems in AFM and describe, where possible, how to overcome them. Knowledge of these limitations is of the utmost importance for correct data acquisition, interpretation, and, ultimately, the embedding of findings into a broad scientific context.

We present a step-by-step approach to identify and address the most common problems associated with atomic force microscopy micro-indentations. We exemplify the emerging problems on native human articular cartilage explants characterized by various degrees of osteoarthritis-driven degeneration.

인간 관절 연골 외식물에 대한 원자력 현미경 기반 미세 압입의 실제 문제 해결

Without a doubt, atomic force microscopy (AFM) is currently one of the most powerful and useful techniques to assess micro and even nano-cues in the ...