3D image analysis of microneedles

Y.Makino1, T.Kurita1, M.Ishibashi2, K.Kobayashi2, H.Hamamoto2, K.Toyohara3 and M.Kiyoki3

1 Tokushima Bunri University, Japan 2 MEDRX Co., Ltd, Japan 3 TEIJIN Limited, Japan

(1)Introduction and Objectives

• Though many pictures of microneedles have been shown, most of them are optical or electron microscopic. Few 3D image analysis has been studied probably due to steep or complex structures of the microneedles. The analysis would contribute to

• Here, non-contact surface profilers such as optical profiler, white-light interferometry or confocal laser microscopy were applied to steep polyglycolic acid microneedles. Possibilities of the optical profiler were particularly focused, as it is a more easier method.

• Successful 3D image analysis would contribute to micrometrology and quality control of a microbeedle and arrays thereof.

(2)Experimentals 【Materials】

1) Microneedles

・PGA microneedles : an array micromolded with polyglycolic acid by TEIJIN (Fig. 1, height : ca. 560μm)

・Hyauronic acid microneedles : an array distributed by SHISEIDO Co., Ltd. (JAPAN) for cosmetic uses (Fig.2, height : ca. 200μm)

・Stainless steel microneedle :

a)rough surface : a microneedle prepared by cutting

(Fig.3 (a), height : ca. 600μm)

b)smooth (polished) surface : a microneedle manufactured by SEIRIN CORPORATION (JAPAN) for acupunctures (Fig.3 (b), height : ca. 600μm)

【Surface profilers】

1)Optical profiler(focus variation method)

Zeta Instruments Z-200 (USA) (Fig.4)

(TEC SCIENCE Co., Ltd (Japan) was appreciated for their assistances.)

2)White-light interferometry

Bruker ContourGT K0 (USA)

(BRUKER AXS Co., Ltd (Japan) was appreciated for their assistances.)

3) Confocal Laser Microscopy

Olympus LEXT OLS4000 (Japan)

(OLYMPUS Co., Ltd (Japan) was appreciated for their assistances.)

【Degital microscope】

KEYENCE VHX 500 (Japan)

【Conditions for 3D image analysis】

1) Optical profiler (focus variation method)

・Spec. of object lenses (with Relay lens (NA: 0.5) and 2/3” CCD)

Magnification 10x 20x 50x

NA 0.30 0.45 0.80

WD (mm) 11.0 3.10 1.00

Field of view (um) 1308*981 654*490 262*196

Vertical resolution (um) 1.50 0.50 0.10

Lateral resolution (um) 1.30 0.651 0.262

2)White-light interferometry

・Spec. of object lenses

Magnification 50x

Field of view (um) 235*176

3) Confocal Laser Microscopy

・Spec. of object lenses

Magnification 10x 20x 50x

NA 0.30 0.60 0.95

WD (mm) 11.0 0.90 0.30

Field of view (um) 2560*2560 1280*1280 640*640

【Metrology of microneedles】

• As for the PGA microneedles, heights, bottom diameters and pitches of the microneedles and flatness of the array floor were measured or observed by the optical profiler Z-200.

• The optical profiler were also applied to detect defects such as tip missing or bending in the microneedles.

• Microneedle(s) made of stainless steel or hyaluronic acid were also analysed by the opticalprofiler.

• 3D analysis of the PGA microneedles by the optical profiler was compared with those by white-light interferometry or confocal laser microscopy.

(3)Results and discussions

• 3D images of the steep PGA microneedles were successfully obtained by the optical profiler Z-200 (Fig.5 and 6). Though confocal laser microscopy gave similar results(Fig.10), white-light interferometry failed to visualize the steep slope(Fig.11).

Though optical profilers are well known, few products gave complete images of the steep slopes. The success of the Z-200 may be due to projection of grids (which is installed after light source) to subjective, which clarifies focused area (Fig. 14).

• Heights, bottom diameters and pitches of the microneedles and flatness of the array floor were successfully measured or observed by the optical profiler Z-200 (Fig. 5 and 6, Table 1).

• The defects such as tip missing or bending in the microneedles were succcessfully detected (Fig.8 and 9, Table 2 )

• Combinations of the each image gave total view of the array (Fig.7).

• As for hyaluronic acid microneedles, which is more transparent and smooth than PGA, the optical profiler failed to give complete image of the steep slopes (data not shown). Confocal laser microscopy, however, gave successful image (Fig. 12).

• As for stainless steel microneedle(s), the image depends on roughness of the surface. Complete image of the steep slope by the profiler was obtained only with rough surface (data not shown). White-light interferometry gave good image for the microneedles with rough and gentle slope (Fig.13).

(4)Conclusions

Optical profiler Z-200 (Zeta Instruments) gave successful 3D image of PGA microneedles with steep slopes. The profiler would contribute to micrometrology and quality controls of the microneedles. Though the technology could not be applied to all kinds of microneedles, it has advantages such as easy handlings and less price over confocal laser microscopy.

Fig.1. Microscopy of a PGA microneedle array

500 um

Fig.2. Microscopy of a hyaluronic acid microneedle array

Fig.3. Microscopy of stainless microneedles

(a) a cutted microneedle with rough surface

(b) a microneedle for acupunctures with smooth surface

Fig.4. Optical profiler Z-200 (Zeta Instruments , USA)

Fig.5. 3D image analysis of the PGA microneedles by the optical profiler (x50)

height

diameter of bottom

Diameter of half height

Fig.6. 3D image analysis of the PGA microneedles by the optical profiler (x10)

Fig.7. 3D image analysis of the PGA microneedles by the optical profiler

( 5 x 7 pictures were combined.)

Table 1. PGA microneedle height determinations

Specific microneedle Degital microscope Optical profiler (Z-200)in an array

x 10 X 50

No.F1 560 587 562

No.F6 560 581 566

No.K1 560 578 563

(μm)

Fig.8. PGA microneedle tip missing detections by the optical profiler

Table 2. Tip missing detections by the Optical profiler

Specific microneedle Degital microscope Optical profilerin an array (x50)

No.D11 150 432

No.E1 250 342

No.F3 270 320

No.F6 270 319

No.G1 280 310

No.H11 240 327

(μm)

Fig.9. PGA microneedle bending detections by the optical profiler

Fig.10. 3D image analysis of the PGA microneedle by confocal laser microscopy (x20)

Fig.11. 3D image analysis of the PGA microneedle by white-light interferometry (x50)

Fig.12. 3D image analysis of the hyaluronic acid microneedles

(a) Optical profiler (x20)

(b) Confocal laser microscopy (x50)

Fig.13. 3D image analysis of the stainless microneedle by white-light interferometry (x50)

Fig.14. Picture of the projected grids on a microneedle.