CN103405215B - A kind of method of intravital mouse retina measuring multiple parameters - Google Patents
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Abstract
The invention discloses a kind of method of intravital mouse retina measuring multiple parameters, the step of the method is as follows: be first fixed on three-dimensional experiment platform by anaesthetizing the mice successfully; Then planoconcave lens, supplementary lens and OCT instrument are assembled; Adjustment three dimensional taest platform makes eyeball of mouse be positioned at the dead ahead of supplementary lens and make scanning retina eye fundus image comparatively clearly appears in OCT instrument display screen; Select high definition one-line scanning pattern, obtain the panretinal faultage image of mice and record the thickness of each layer on Mouse Retina; Select retina volume mode, mice optic disc place is aimed in the center of cross scanning line, records average thickness and the volume of Mouse Retina 9 sub regions, and the form of three-dimensional reconstruction Mouse Retina; Finally select optic disc scan pattern, obtain the continuous thickness data of retinal nerve fibre layer.The present invention has simple, favorable repeatability, result accurately, not by the feature of external environmental interference, suitablely to promote the use of.
Description
Technical field
The present invention relates to Small Rodents living animal retina field of measuring technique, specifically a kind of optical coherence tomography carries out the method for intravital mouse retina measuring multiple parameters.
Background technology
Mice, as a kind of laboratory animal highly similar with human genotype, is widely used in medical research, so same in ophthalmology basic research.Therefore, a kind of Research of Animal Model for Study of Mouse Retina measuring technique to ophthalmic diseases of mature and reliable has great significance.The retina of current mice is measured and extract the observation of eyeball capable Histological section after being put to death mice.But eyeball of mouse is too little, in slicing processes, ensure that the integrity of tissue is the difficulty work needing protracted experience to accumulate; Secondly, the retinopathy of mice is very trickle, the situation of omitting when often there is section; Finally, in order to the research of longitudinal 2 observation in time needs to sacrifice a large amount of mices, not only wasted funds but also be unfavorable for the requirement of animal protection.Therefore mice live body retina measuring method has the Practical significance of height.
Optical coherence tomography (Optical Coherence Tomography, OCT), it is a kind of diagnostic techniques being applied to clinical ophthalmology in recent years, there is the advantages such as not damaged, cell grade high-resolution, fast imaging and precise quantitative measurement, use OCT can solve the problem of mice live body retina research well.
Summary of the invention
The object of the invention is for prior art Problems existing, provide a kind of simple, measurement parameter is many, measurement parameter is wide and measure the method for conveniently intravital mouse retina measuring multiple parameters.
The object of the invention is to solve by the following technical programs:
A method for intravital mouse retina measuring multiple parameters, is characterized in that the step of the method is as follows:
A () is fixed on three-dimensional experiment platform by anaesthetizing the mice successfully;
B () mydriatic falls apart after large mice pupil and forms with coverslip and concavees lens the planoconcave lens being positioned at eyeball of mouse front surface;
C () installs supplementary lens on the camera lens of OCT instrument, adjustment three dimensional taest platform makes eyeball of mouse be positioned at the dead ahead of supplementary lens, and by far away and scanning retina eye fundus image comparatively clearly appears in the display screen of near until OCT instrument;
D () selects high definition one-line scanning pattern, by scanning line center aligned mice optic disc place, obtain the panretinal faultage image of mice and record the thickness of each layer on Mouse Retina;
E () then selects retina volume mode, mice optic disc place is aimed in the center of cross scanning line, records with the average thickness of the optic disc center Mouse Retina that is the center of circle and volume, and the form of three-dimensional reconstruction Mouse Retina;
F () finally selects optic disc scan pattern, obtain with the continuous thickness data of the retinal nerve fibre layer of optic disc center certain area that is the center of circle.
During high definition one-line scanning pattern in described step (d), on Mouse Retina, the thickness of each layer is recorded by the survey tool on OCT instrument.
During high definition one-line scanning pattern in described step (d), the scale conversion equation that OCT instrument is imaged on Mouse Retina horizontal plane direction is: actual value=0.224*OCT measured value+72.61, and wherein the unit of actual value, OCT measured value and constant 72.61 is μm; Then there is not amplification in the measurement of OCT instrument on retina vertical meridian direction.
Retina volume mode in described step (e) selects macula retinae volume scan pattern.
The scale conversion equation that the retina scanning area that retina volume mode in described step (e) obtains is imaged on Mouse Retina horizontal plane direction according to the sweep diameter of OCT instrument and OCT instrument is determined.
Retina scanning area in described step (e) be with optic disc center be the center of circle, diameter is respectively in the concentric circular of the central area of 1 ㎜, 3 ㎜ and 6 ㎜ (the corresponding diameter on Mouse Retina be 0.297,0.745 and 1.417mm), each four quadrants Mouse Retina average thickness of totally 9 sub regions and volume up and down.
The scale conversion equation that the layer of fibers of optic nerve of retina area that optic disc scan pattern in described step (f) obtains is imaged on Mouse Retina horizontal plane direction according to the sweep diameter of OCT instrument and OCT instrument is determined.
The present invention has the following advantages compared to existing technology:
The present invention adopts the optical instruments such as clinical OCT instrument to measure intravital mouse retina, and resolution is high, distinguishable Mouse Retina tomography more than ten layers of measuring, substantially corresponding with Histological section viewed layer of structure; Measurement effect can replace even beyond tradition section measuring method, is conducive to the progression of longitudinal 2 observation Mouse Retina pathological changes, decreases the quantity of required laboratory animal simultaneously.
The present invention is easy to location and quantitative check, this mice live body retina measuring method can obtain scanning laser eye fundus image and OCT tomoscan image simultaneously, synchronous optical fundus and OCT image make research worker can obtain the faultage image of Mouse Retina any part, only need simply by the position of mouse drag scanning line, the angle of definition laser scanning line.
The present invention can to Mouse Retina fast, batch measuring multiple parameters, if retina volume mode is by single pass (about 1 second), can obtain mice centered by optic disc, diameter is respectively in the concentric circular of the central area of 1 ㎜, 3 ㎜ and 6 ㎜, the average thickness of each four quadrants totally 9 sub regions up and down, obtains amphiblestroid form and carries out three-dimensional reconstruction (altogether the automatic measurement of or) at 512 × 128 at 200 × 200; Optic disc scan pattern simultaneously can also go out the indexs such as Mouse Retina nerve fibre layer average thickness by automatic measurement & calculation, these fast, high flux measure be all traditional histological section measurement incomparable.
Method of the present invention is simple, favorable repeatability, and result is accurate, not by external environmental interference, can obtain Mouse Retina faultage image clearly and measure for mice live body retina, effectively reduce the quantity of experiment mice, save reasearch funds; The retinal structure form of viviperception ophthalmic diseases animal model is played an important role, is suitable for promoting the use of.
Accompanying drawing explanation
The apparatus structure schematic diagram that accompanying drawing 1 adopts for inspection method of the present invention;
Measurement result gained image when accompanying drawing 2 is high definition one-line scanning pattern of the present invention on direction, retinal levels face;
Accompanying drawing 3 is the Histological section gained image corresponding with accompanying drawing 2;
OCT measured value when accompanying drawing 4 is high definition one-line scanning pattern of the present invention on direction, retinal levels face and the loose point of paving sheet measured value and matched curve figure;
Accompanying drawing 5 is the Mouse Retina faultage image of high definition one-line scanning pattern gained of the present invention;
The OCT measured value that when accompanying drawing 6 is high definition one-line scanning pattern of the present invention, thickness measure records and the loose point of section measured value and matched curve figure;
Accompanying drawing 7 is that retina volume mode of the present invention measures the Mouse Retina nine sub regions average thickness of gained and general area thickness and volumetric values, wherein Fig. 7 A represents mice optical fundus OCT scintigram, Fig. 7 B represents the retina average thickness values of nine sub regions centered by mice optic disc, and Fig. 7 C represents retina volume in subarea, the center thickness of Mouse Retina, scanning area and average thickness (ILM-RPE distance);
Accompanying drawing 8 is image and the numerical information of the Mouse Retina nerve fibre layer (RNFL) of optic disc volume scan pattern gained of the present invention, wherein Fig. 8 A is that (OD represents left eye to mice eyes OCT optical fundus scanogram, OS represents right eye, border circular areas is the data measurement sites of RNFL), Fig. 8 B is that (OD represents left eye for the RNFL average thickness of mice eyes measuring position, OS represents right eye, unit is μm), 8C is that (OD represents left eye to the RNFL average thickness represented with four quadrants at the bottom of mice eyes, OS represents right eye, unit is μm), 8D is that (OD represents left eye to the RNFL thickness represented with 12 hours at the bottom of mice eyes, OS represents right eye, unit is μm).
Wherein: 1:OCT instrument; 2: supplementary lens; 3: planoconcave lens; 4: coverslip; 5: concavees lens; 6: three dimensional taest platform; 7: eyeball of mouse.
GCL: ganglion cell layer of retina; IPL: inner plexiform layer of retina; INL: inner nuclear layer of retina; OPL: outer plexiform layer of retina; ONL: outer nuclear layer; IS-OS: retinal light injury photoreceptor is inside and outside to be connected; RPE: layer of retina,pigment epithelium; ILM-RPE: inner limiting membrane-layer of retina,pigment epithelium; RNFL: retinal nerve fibre layer.
Detailed description of the invention
Below in conjunction with accompanying drawing and embodiment, the present invention is further illustrated.
As shown in Figure 1: a kind of method of intravital mouse retina measuring multiple parameters, the step of the method is as follows: the anesthetis that (a) adopts the mixed solution of ketamine and xylazine to make, according to 0.2-0.5ml/20g body weight intraperitoneal injection of anesthesia mice, is anaesthetized successfully and is fixed on by mice on three-dimensional experiment platform 6; The mydriatic that Tropicamide and Phenylephrine b () then adopts normal saline dilution after is made falls apart after large mice pupil and concavees lens 5 is covered eyeball of mouse 7 surface, gets coverslip 4 and is placed on concavees lens 5 and makes coverslip 4 and concavees lens 5 form the planoconcave lens 3 being positioned at eyeball of mouse 7 front surface; C () installs supplementary lens 2 on the camera lens of OCT instrument 1, adjustment three dimensional taest platform 6 makes eyeball of mouse 7 be positioned at the dead ahead of supplementary lens 2, and by far away and scanning retina eye fundus image comparatively clearly appears in the display screen of near until OCT instrument 1; D () selects high definition one-line scanning pattern, by scanning line center aligned mice optic disc place, obtain the panretinal faultage image of mice, and recorded the thickness of each layer on Mouse Retina by the survey tool on OCT instrument 1; E () then selects retina volume mode, mice optic disc place is aimed in the center of cross scanning line, records average thickness and the volume of Mouse Retina 9 sub regions, and the form of three-dimensional reconstruction Mouse Retina; F () finally selects optic disc scan pattern, can obtain the continuous thickness data of retinal nerve fibre layer.In above-mentioned steps, during high definition one-line scanning pattern, the measured value of OCT instrument 1 and optical microscope measuring value height correlation, there is not amplification in the measurement that OCT instrument 1 is imaged on Mouse Retina vertical meridian direction; And the measurement that OCT instrument 1 is imaged on Mouse Retina horizontal plane direction exists significant amplification, there is fixing conversion in this amplification in measuring system of the present invention, the scale conversion equation that OCT instrument 1 is imaged on Mouse Retina horizontal plane direction is: actual value=0.224*OCT apparatus measuring value+72.61, and wherein the unit of actual value, OCT apparatus measuring value and constant 72.61 is μm.The continuous thickness data of the Mouse Retina nerve fibre layer obtained when the average thickness in Mouse Retina 9 regions obtained during retina volume mode in addition and volume and optic disc scan pattern, the scale conversion equation that the actual size of optic disc is imaged on Mouse Retina horizontal plane direction according to the sweep diameter of OCT instrument 1 and OCT instrument 1 is determined; Retina scanning area in addition during retina volume mode refer to optic disc center be the center of circle, diameter is respectively in the concentric circular of the central area of 1 ㎜, 3 ㎜ and 6 ㎜, each four quadrants Mouse Retina average thickness of totally 9 sub regions and volume up and down; Be respectively the central area of 1 ㎜, the concentric circular of 3 ㎜ and 6 ㎜ relative to diameter on optic disc, then the corresponding diameter on Mouse Retina is respectively 0.297,0.745 and 1.417mm.
After adding the optical imagery instruments such as supplementary lens for checking in addition, OCT instrument 1 imaging is erect image or inverted image, and before checking, we are to mice row Fundus laser, above its optic disc temporo, form a laser spot; Use the 1 pair of Mouse Retina imaging of OCT instrument subsequently, viewed laser spot position, still above optic disc temporo, draws thus: by after adding optical instrument such as supplementary lens 2 grade become image to remain erect image.
Embodiment one
Select C57BL mouse, note before checking checking that whether mice refracting media is transparent; Adopt clinical modal OCT instrument 1:Zeiss Cirrus HD-OCT 4000 or 400 simultaneously.The anesthetis that the mixed solution getting ketamine and xylazine is made is according to 0.2ml/20g body weight intraperitoneal injection of anesthesia mice, anaesthetize successfully and mice row Fundus laser light is coagulated, a laser spot (diameter 50 μm, punctures Bruch film) is formed at Mouse Retina random site.Subsequently mice is fixed on three-dimensional experiment platform 6; Then the mydriatic adopting the Tropicamide and Phenylephrine after normal saline dilution to make falls apart after large mice pupil and medical hyaluronic acid viscoelastic agent (as concavees lens 5) is covered eyeball of mouse 7 surface, gets coverslip 4 and is placed in the planoconcave lens 3 hyaluronic acid viscoelastic agent being formed and is positioned at eyeball of mouse 7 front surface; Be before the Volk Superfield NC supplementary lens 2 of 90D is fixedly installed on the camera lens of OCT instrument 1 by a diopter, adjustment three dimensional taest platform 6 makes eyeball of mouse 7 be positioned at dead ahead about the 5cm of supplementary lens 2, and by far away and near until there is eye fundus image; The position of adjustment three dimensional taest platform 6 until OCT instrument 1 display screen on there is scanning retina eye fundus image comparatively clearly.Select the high definition one-line scanning pattern of OCT instrument 1 to Mouse Retina imaging, adjustment scanning line angle is located on the line of optic disc and laser spot, records the distance (as shown in Figure 2) between optic disc to laser spot under the imaging of OCT instrument.Put to death mice, get the capable overall inner nuclear layer retina of its eyeball, measure the distance (as shown in Figure 3) drawn between optic disc to laser spot under optical microscope, totally 8 mices, every each eyeball of mice has a laser spot.
Adopt SPSS19.0 statistical software, OCT instrument 1 data measured and inner nuclear layer retina the data obtained are carried out correlation and regression analysis, OCT measured value during high definition one-line scanning pattern and the loose point of section measured value and matched curve figure as shown in Figure 4, square R of regression analysis neutral line correlation coefficient
2=0.887.Finally obtain the scale conversion equation on OCT instrument 1 imaging Mouse Retina horizontal plane direction (x-axis, y-axis): the unit of actual value=0.224*OCT value+72.61(wherein actual value, OCT apparatus measuring value and constant 72.61 is μm).Paving sheet measure actual value and the regression analysis of OCT measured value as shown in the table:
In above process, due to the physiological characteristics of mice, eye birth prevalence is high, and refracting media easily affects by medicine, pressure, hydropenia and environment etc. and muddiness occurs very soon, so need before experiment to check eye conditions, avoid selecting the animal of refractive media difference in order to avoid affect result; Need careful operation during experiment, namely adopt viscoelastic agent eye dripping to add successfully to cover with coverslip 4, namely can form a planoconcave lens 3 and participate in optical imagery, the evaporation of mice eye moisture can be reduced again once anaesthetize, prevent its refractive media muddy.
Embodiment two
Select C57BL mouse, note before checking checking that whether mice refracting media is transparent; Adopt clinical modal OCT instrument 1:Zeiss Cirrus HD-OCT 4000 or 400 simultaneously.The anesthetis that the mixed solution getting ketamine and xylazine is made, according to 0.2ml/20g body weight intraperitoneal injection of anesthesia mice, is anaesthetized successfully and is fixed on by mice on three-dimensional experiment platform 6; Then the mydriatic adopting the Tropicamide and Phenylephrine after normal saline dilution to make falls apart after large mice pupil and medical hyaluronic acid viscoelastic agent (as concavees lens 5) is covered eyeball of mouse 7 surface, gets coverslip 4 and is placed in the planoconcave lens 3 hyaluronic acid viscoelastic agent being formed and is positioned at eyeball of mouse 7 front surface; Be before the Volk Superfield NC supplementary lens 2 of 90D is fixedly installed on the camera lens of OCT instrument 1 by a diopter, adjustment three dimensional taest platform 6 makes eyeball of mouse 7 be positioned at dead ahead about the 5cm of supplementary lens 2, and by far away and near until there is eye fundus image; The position of adjustment three dimensional taest platform 6 until OCT instrument 1 display screen on there is scanning retina eye fundus image comparatively clearly.Select high definition one-line scanning pattern, by scanning line center aligned mice optic disc place, according to the angle of the difference adjustment scanning line of required look-out station, to obtain the panretinal faultage image of mice (as shown in Figure 5).The full thickness of Mouse Retina different parts and the thickness of this position inner nuclear layer, outer nuclear layer is measured with the survey tool that OCT instrument is subsidiary.
Put to death the mice of OCT instrument data measured, get the capable Histological section of its eyeball.Above Histological section chooses respectively, below, endocanthion, outer canthus four direction distance optic nerve hit exactly the position measurement retina full thickness of 200 μm, 300 μm, 400 μm and 500 μm and inside and outside stratum nucleare thickness, according to the scale conversion equation of trying to achieve in embodiment one, OCT instrument is measured the retina full thickness at position corresponding with Histological section and inside and outside stratum nucleare thickness.The OCT measured value that during high definition one-line scanning pattern, thickness measure records and the loose point of section measured value and matched curve figure are as shown in Figure 6, OCT data measured and section data measured are compared, adopt SPSS19.0 statistical software to carry out correlation and regression analysis, analysis result is as shown in the table:
Can be obtained by statistical analysis: the ratio of section value and OCT value is approximately equal to 1.Because eyeball of mouse needs through the step such as fixing of dewatering in section statining process, can there is shrinkage to a certain degree in its retina, in addition certain measurement error after dewatering, so section measured value is more smaller than OCT apparatus measuring value.Can be known by the operation principle of OCT instrument, what OCT instrument was measured is the luminous reflectance signal of laser in retina different levels, time difference according to luminous reflectance signal carrys out detect thickness, the imaging of OCT image in retina z-axis can not cause due to additional optical components and zoom in or out in theory, and this is also consistent with our measurement result.Can think: OCT instrument is imaged in Mouse Retina z-axis does not exist amplification phenomenon, and when the imaging of OCT instrument and Histological section, the measurement result of optical microscope gained is basically identical.
In above process, due to the physiological characteristics of mice, eye birth prevalence is high, and refracting media easily affects by medicine, pressure, hydropenia and environment etc. and muddiness occurs very soon, so need before experiment to check eye conditions, avoid selecting the animal of refractive media difference in order to avoid affect result; Need careful operation during experiment, namely adopt viscoelastic agent eye dripping to add successfully to cover with coverslip 4, namely can form a planoconcave lens 3 and participate in optical imagery, the evaporation of mice eye moisture can be reduced again once anaesthetize, prevent its refractive media muddy.
Embodiment three
Select C57BL mouse, note before checking checking that whether mice refracting media is transparent; Adopt clinical modal OCT instrument 1:Zeiss Cirrus HD-OCT 4000 or 400 simultaneously.The anesthetis that the mixed solution getting ketamine and xylazine is made, according to 0.2ml/20g body weight intraperitoneal injection of anesthesia mice, is anaesthetized successfully and is fixed on by mice on three-dimensional experiment platform 6; Then the mydriatic adopting the Tropicamide and Phenylephrine after normal saline dilution to make falls apart after large mice pupil and medical hyaluronic acid viscoelastic agent (as concavees lens 5) is covered eyeball of mouse 7 surface, gets coverslip 4 and is placed in the planoconcave lens 3 hyaluronic acid viscoelastic agent being formed and is positioned at eyeball of mouse 7 front surface; Be before the Volk Superfield NC supplementary lens 2 of 90D is fixedly installed on the camera lens of OCT instrument 1 by a diopter, adjustment three dimensional taest platform 6 makes eyeball of mouse 7 be positioned at dead ahead about the 5cm of supplementary lens 2, and by far away and near until there is eye fundus image; The position of adjustment three dimensional taest platform 6 until OCT instrument 1 display screen on there is scanning retina eye fundus image comparatively clearly.Select retina volume mode, the center of cross scanning line is aimed at mice optic disc centre, by this pattern draw refer to optic disc center be the center of circle, diameter is respectively in the concentric circular of the central area of 1 ㎜, 3 ㎜ and 6 ㎜, each four quadrants Mouse Retina average thickness of totally 9 sub regions and volume up and down, be respectively the central area of 1 ㎜, the concentric circular of 3 ㎜ and 6 ㎜ relative to diameter on optic disc, then the corresponding diameter on Mouse Retina is respectively 0.297,0.745 and 1.417mm; And can the panretinal form of three-dimensional reconstruction mice (as shown in Figure 7).During OCT instrument illustrates, the sweep diameter of this scan pattern is 6mm, amplifies to some extent after Additional optical lens, and the scale conversion equation substituting into gained in example 1 is known, and the sweep diameter on actual Mouse Retina is 1.417mm.
Embodiment four
Select C57BL mouse, note before checking checking that whether mice refracting media is transparent; Adopt clinical modal OCT instrument 1:Zeiss Cirrus HD-OCT 4000 or 400 simultaneously.The anesthetis that the mixed solution getting ketamine and xylazine is made, according to 0.2ml/20g body weight intraperitoneal injection of anesthesia mice, is anaesthetized successfully and is fixed on by mice on three-dimensional experiment platform 6; Then the mydriatic adopting the Tropicamide and Phenylephrine after normal saline dilution to make falls apart after large mice pupil and medical hyaluronic acid viscoelastic agent (as concavees lens 5) is covered eyeball of mouse 7 surface, gets coverslip 4 and is placed in the planoconcave lens 3 hyaluronic acid viscoelastic agent being formed and is positioned at eyeball of mouse 7 front surface; Be before the Volk Superfield NC supplementary lens 2 of 90D is fixedly installed on the camera lens of OCT instrument 1 by a diopter, adjustment three dimensional taest platform 6 makes eyeball of mouse 7 be positioned at dead ahead about the 5cm of supplementary lens 2, and by far away and near until there is eye fundus image; The position of adjustment three dimensional taest platform 6 until OCT instrument 1 display screen on there is scanning retina eye fundus image comparatively clearly.Select optic disc volume scan pattern, the analytical tool that OCT instrument 1 carries can obtain with retinal nerve fibre layer (the Retinal Nerve Fiber Layer on the 0.848mm diameter line that is the center of circle of mice optic disc center, RNFL) thickness scintigram, and with 12 hours region display RNFL varied in thickness (as shown in Figure 8).The circle diameter that during OCT instrument 1 illustrates, this scan pattern RNFL measures is 3.46mm, amplifies to some extent after Additional optical lens, and the scale conversion equation substituting into gained in example 1 is known, and it is 0.848mm that actual Mouse Retina RNFL measures diameter.
By the measurement of above embodiment to Mouse Retina, OCT instrument 1 is imaged on the upper existence in Mouse Retina horizontal plane direction (x-axis, y-axis) and amplifies phenomenon, and there is not amplification phenomenon in z-axis.This mice live body retina measuring method can replace the application of Histological section in Mouse Retina is measured.In addition, some process of measurement instruments that OCT instrument 1 carries all can be used for the somatometry of mice, such as, can record the average thickness of Mouse Retina, mice RNFL thickness etc. under retina volume mode, these data are all that routine histologic section cannot accurately obtain, and then can run away with result by this mice live body retina measuring method.Should be noted, there is amplification phenomenon to Mouse Retina plane (x-axis, y-axis) in optical instrument such as OCT instrument 1 grade, need obtain correspondence position when measuring by converting.
Method of the present invention is simple, favorable repeatability, and result is accurate, not by external environmental interference, can obtain Mouse Retina faultage image clearly and measure for mice live body retina, effectively reduce the quantity of experiment mice, save reasearch funds; The retinal structure form of viviperception ophthalmic diseases animal model is played an important role, is suitable for promoting the use of.
Above embodiment is only and technological thought of the present invention is described, can not limit protection scope of the present invention with this, every technological thought proposed according to the present invention, and any change that technical scheme basis is done, all falls within scope; The technology that the present invention does not relate to all is realized by prior art.
Claims (6)
1. a method for intravital mouse retina measuring multiple parameters, is characterized in that the step of the method is as follows:
A () is fixed on three-dimensional experiment platform by anaesthetizing the mice successfully;
B () mydriatic falls apart after large mice pupil and forms with coverslip and concavees lens the planoconcave lens being positioned at eyeball of mouse front surface;
C () installs supplementary lens on the camera lens of OCT instrument, adjustment three dimensional taest platform makes eyeball of mouse be positioned at the dead ahead of supplementary lens, and by far away and scanning retina eye fundus image comparatively clearly appears in the display screen of near until OCT instrument;
D () selects high definition one-line scanning pattern, by scanning line center aligned mice optic disc place, obtain the panretinal faultage image of mice and record the thickness of each layer on Mouse Retina;
E () then selects retina volume mode, mice optic disc place is aimed in the center of cross scanning line, records with the average thickness of the optic disc center Mouse Retina that is the center of circle and volume, and the form of three-dimensional reconstruction Mouse Retina;
F () finally selects optic disc scan pattern, obtain with the continuous thickness data of the retinal nerve fibre layer of optic disc center certain area that is the center of circle;
During high definition one-line scanning pattern in described step (d), the scale conversion equation that OCT instrument is imaged on Mouse Retina horizontal plane direction is: actual value=0.224*OCT measured value+72.61, and wherein the unit of actual value, OCT measured value and constant 72.61 is μm.
2. the method for intravital mouse retina measuring multiple parameters according to claim 1, when it is characterized in that the high definition one-line scanning pattern in described step (d), on Mouse Retina, the thickness of each layer is recorded by the survey tool on OCT instrument.
3. the method for intravital mouse retina measuring multiple parameters according to claim 1, is characterized in that the retina volume mode in described step (e) selects macula retinae volume scan pattern.
4. the method for intravital mouse retina measuring multiple parameters according to claim 1, is characterized in that the scale conversion equation that the retina scanning area that the retina volume mode in described step (e) obtains is imaged on Mouse Retina horizontal plane direction according to the sweep diameter of OCT instrument and OCT instrument is determined.
5. the method for the intravital mouse retina measuring multiple parameters according to claim 1 or 4, the retina scanning area that it is characterized in that in described step (e) be with optic disc center be the center of circle, diameter is respectively in the concentric circular of the central area of 1 ㎜, 3 ㎜ and 6 ㎜, each four quadrants totally 9 sub regions up and down.
6. the method for intravital mouse retina measuring multiple parameters according to claim 1, is characterized in that the scale conversion equation that the layer of fibers of optic nerve of retina area that the optic disc scan pattern in described step (f) obtains is imaged on Mouse Retina horizontal plane direction according to the sweep diameter of OCT instrument and OCT instrument is determined.
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| CN103211576A (en) * | 2013-04-22 | 2013-07-24 | 江苏省人民医院 | Wide-angle optical imaging system for mouse retina optical coherence tomography (OCT) inspecting and inspecting method with wide-angle optical imaging system |
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| CN103211576A (en) * | 2013-04-22 | 2013-07-24 | 江苏省人民医院 | Wide-angle optical imaging system for mouse retina optical coherence tomography (OCT) inspecting and inspecting method with wide-angle optical imaging system |
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