yong-li pan, dec. 28, 2011 ( biomaterial chemistry c ourse)
DESCRIPTION
Invasive candidiasis ( 念珠菌病 ). Invasive aspergillosis ( 麴菌病 ). Diagnosis of infectious disease and pathogens based on protein biomarkers of MALDI-TOF mass spectrometry. Discovery/Development of Biomarker panels by Systems Biology. - PowerPoint PPT PresentationTRANSCRIPT
1
Proteomics
Diagnosis of infectious disease and pathogens based on protein biomarkers of MALDI-TOF mass spectrometryYong-Li Pan, Dec. 28, 2011 (Biomaterial
Chemistry Course)Invasive candidiasis (念珠菌病 ) Invasive aspergillosis (麴菌病
)
Genomics; Functional genomics Metabolomics
0.0
0.5
1.0
1.54x10
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
0.00
0.25
0.50
0.75
1.00
1.254x10
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
Oligonucleotide arraycDNA microarray
ESI mass spectrometryMALDI mass spectrometry Normal Raman spectroscopy
SERS Raman spectroscopy
(Riboflavin, Pigment, etc.)
Discovery/Development of Biomarker panels by Systems Biology
1
Corneal ulcerationInfectious corneal ulcer
Infectious corneal ulcer is more centrally located, more discharge and greater vision-threatening than noninfectious.
Noninfectious corneal ulcer
A typical presentation of noninfectious corneal ulcer is more peripheral distribution, a clear zone between lesion and adjacent limbus.
However, sometimes they are difficult to differentiate in several cases without typical presentation.
3
Species identification of lethal Aspergillus by detecting single conidiospores base on Raman spectroscopy
Stage I of A. fumigatus Stage II of A. fumigatus
2nd day
On SDA plate, 30~35 ℃Sporulation
1st day
Single spores acquired before the significant sporulation on the whole agar plate
Definition of Biomarkers• Medical or clinical biomarker
– FDA: A characteristic that is objectively measured and evaluated as an indicator of normal biologic processes, pathogenic processes, or pharmacologic responses to a therapeutic intervention.
• Environmental or pollution biomarker– Microorganisms ( 指標微生物 )– For monitoring pollution
• Drug target marker– For target therapy (neutralization/blocking by humanized
antibody)– With drug releasing or photothermal mateial/device 4
Molecular types of biomarkers
• Proteins: proteomics, mass spectrometry
• mRNA expression profiling: cDNA microarray, qPCR
• Ribosomal DNA or SNP (single nucleotide polymorphisms)
• Small molecules: metabonomics
5
Clinical application of biomarker• Screening (CEA, AFP, CA125, PSA)• Diagnosis (Tumor, infectious disease,
cardiovascular disease, etc.)• Monitoring treatment• Detection of recurrence• Prognosis• Personalized medicine 個人化醫療
(pharmacogenomics and proteomics/molecular diagnostics)
6
Types of tumor markers• Circulating tumor markers
–Enzymes, serum proteins and hormones–Associations with metastases–Prognostic factors–Monoclonal-defined tumor markers–Mucin and blood group substances–Nonspecific tumor markers–Urinary and CSF tumor markers
• Cellular tumor marker7
Discovery/development and validation of biomarkers by biosensing technology
8
ʳ * A16-1800a\0_A16\1\1SLin, "Baseline subt."
0
2
4
6
4x10
Intens. [a.u.]
ʳ
* C18-1800a\0_C18\1\1SLin, "Baseline subt."
0
2
4
6
84x10
Intens. [a.u.]
ʳ
* 7-P aeruginosa 10944-blood culture\0_B7\1\1SLin, "Baseline subt."
0
1
2
3
4x10
Intens. [a.u.]
5000 10000 15000 20000 25000 30000 35000m/z
600 700 800 900 1000 1100 1200 1300 1400 1500 1600 1700 1800
Raman shift (cm-1)
abcdefgh
Proteomics/mass spectrometry
Biochemical component/Raman spectroscopy
Cell physiology/dielectrophresis
9
Identification by conventional methods based on culture for checking
Workflow of developing reliable methods for rapidly screening lethal fungal species based on DNA microarray, Raman spectral and MALDI-TOF mass biosensing technologies
Establishment of reliable diagnostic criteria
Diagnostic performance evaluation
Optimization of rapid microbial detection methods
Statistical calculation and interpretation of experimental dataDiscussions
Characterization of different species
Optimization of rapid and cheap microbial preparation methods
Collection of target specimens according to purpose
Application in clinical microbial laboratories
Poor diagnostic ability
Excellent diagnostic ability and reliability
(short-term culture)
n=5~8 (3 SD)
n=46 (3 SD)
Validation of biomarkers
Detecting valid biomarkers by probes immobilized on biosensors
10
Detection of Tear Proteins Using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry
Drift tube (Vacuum system)
Ion
Det
ecto
r(ti
me
to m
/z)
Acceleration
Coc
ryst
aliz
atio
nof
pr
otei
n an
d m
atrix
+
+
Soft ionization Flight (Constant velocity)
+ + ++m1, v1m2, v2m3, v3
(m3>m2>m1; v1>v2>v3)(Energy: zeV)
Lens
A B
DC
Isolation/CultivationCocrystalization with matrix
Soft ionization
Contact lens solution
Bottom layer methodTear fluidThe early stage of keratitis
12
Different ion sources for mass spectrometry
• Electro Spray Ionization (ESI)– Easily coupling with other
chromatographic and separation technologies (LC-MS) on line
– Without sample preparation– Small m/z ratio signal (<1000 m/z)
• Matrix-Assisted Laser Desorption Ionization (MALDI-TOF)– Sample preparation with matrix
(off line, dried droplet)– Large m/z ratio signal and range
(1000~>10000 m/z)
氧化
還原
小分子 電場
多電荷
表面張力
-
+ Ionization
Spray
毛細管液體
(陽極 )
(陰極 )
13
MALDI: Matrix Assisted Laser Desorption Ionization
hn
Laser
1. Sample (A) is mixed with excess matrix (M) and dried on a MALDI plate.
2. Laser flash de-ionizes matrix molecules (acid).
3. Sample molecules are ionized by proton transfer from matrix:
MH+ + A M + AH+.
AH+
+20 kV
Variable Ground Grid Grid
Sample plate
Sandler Mass Spectrometry User’s Group, UCSF, US
大分子
LaserIonization
Desorption
電場
Cocrystalization
固體
14
Time-of-flight mass analyzer
+
+
+
+
Ion SourceDrift region (flight tube)
dete
ctor
V
• Ions are formed in pulses.• Small ions reach the detector before large ones.• Measures the time for ions to reach the detector.
zeV2
mv 2
zeV2
mv 2
Sandler Mass Spectrometry User’s Group, UCSF, US
ESIMALDI
無電場 ( 等速飛行 )電場加速
15
Voyager-DE STR MALDI-TOF MS
Camera
Laser
Sample plate
Pumping Pumping
Timed ion selector Reflector
Linear detector
Extractiongrids
ReflectordetectorAttenuator
Prism
Collision cell
Sandler Mass Spectrometry User’s Group, UCSF, US
16
Bruker Daltonics MALDI-TOF mass spectrometry
Desorption and ionization by UV Laser (355 nm ND-YAG Laser)
Co-crystalization of matrix and protein
Steel target plateAutoflex, Bruker
Autoflex, Bruker
17
History of MALDI TOF MS applied in medical microbiology
Piseth Seng, et. al., Clinical Infectious Diseases 2009; 49:543–51
Nobel prize in chemistry (Koichi Tanaka, 田中耕一 , and J.B. Fenn, 芬恩 )
First soft ionization technique for high mass molecules (1987)
First MALDI TOF MS of filament fungal spores (2000)
MALDI TOF MS applied in pathogenic fungi
John B. Fenn :將 ESI (Electro Spray Ionization) 電灑游離法應用在蛋白質 . 生化大分子的分析上Koichi Tanaka :以 MALDI (Matrix Assisted Laser Desorption Ionization) 基質輔助雷射脫附法來分析生化大分子
First MALDI TOF MS of whole fungal cell (2001)
18
Application of MALDI TOF MS in rapid detection of bacterial isolation
Traditional methods
MALDI TOF MS
Isolation from blood culturePiseth Seng, et. al., Clinical Infectious Diseases 2009; 49:543–51
19
Delays, costs, and Level of training for isolate identification methods
Piseth Seng, et. al., Clinical Infectious Diseases 2009; 49:543–51
20
Basic workflow of mass spectrometry
http://pubs.niaaa.nih.gov/publications/arh26-3/images/israel1.gif
Separated, degraded peptide or lysed microorganisms
ESI or MALDI
TOF-MS or FT-MS
Mass spectral patternAmino acid sequencing
Ion detection
ʳ * Candida parapsilosis_BCRC-21544\0_K5\1\1Lin, "Baseline subt."
0200040006000
Intens. [
a.u.]
ʳ
* Candida lusitaniae_CBS-7270\0_C18\1\1Lin, "Baseline subt."
0200040006000
Intens. [
a.u.]
ʳ
* Candida glabrata_CBS-2175\0_C20\1\1Lin, "Baseline subt."
0
2000
4000
Intens. [
a.u.]
ʳ
* Candida krusei_BCRC-21720\0_C16\1\1Lin, "Baseline subt."
0200040006000
Intens. [
a.u.]
ʳ
* Candida albicans_BCRC-20519\0_C3\1\1Lin, "Baseline subt."
0
2000
4000
Intens. [
a.u.]
ʳ
* Candida tropicalis_BCRC-560\0_A14\1\1Lin, "Baseline subt."
0200040006000
Intens. [
a.u.]
5000 10000 15000 20000 25000 30000m/z
20http://www.peptide2.com/peptide/Wikipedia_Mass_Spectrometry_files/Ms_block_schematic.gif
21
ʳ * Candida parapsilosis_BCRC-21544\0_K5\1\1Lin, "Baseline subt."
0200040006000
Inten
s. [a.
u.]ʳ
* Candida lusitaniae_CBS-7270\0_C18\1\1Lin, "Baseline subt."
0200040006000
Inten
s. [a.
u.]
ʳ
* Candida glabrata_CBS-2175\0_C20\1\1Lin, "Baseline subt."
0
2000
4000
Inten
s. [a.
u.]
ʳ
* Candida krusei_BCRC-21720\0_C16\1\1Lin, "Baseline subt."
0200040006000
Inten
s. [a.
u.]
ʳ
* Candida albicans_BCRC-20519\0_C3\1\1Lin, "Baseline subt."
0
2000
4000
Inten
s. [a.
u.]
ʳ
* Candida tropicalis_BCRC-560\0_A14\1\1Lin, "Baseline subt."
0200040006000
Inten
s. [a.
u.]
5000 10000 15000 20000 25000 30000m/z
Homogenization in 5% formic acid and 35% ACN
Disrupt, lyse cells and extract proteins
100 for 5 mins℃Wet heating inactivation
Bio-safety LabWash, centrifuge,
freeze drying
Dried dropletSinapic acid cocrystalization
UV Laser (337 nm)
Beavis, R. C.; Chait, B. T. Rapid Commun. Mass Spectrom. 1989, 3, 432-435.
Common Standard Lab
3. Discriminatory ability analysis(cluster analysis and peak comparison)
Safe transportation and experiment
21
2. MALDI TOF MS experiment
1. Microorganism inactivation
Living yeast Dead yeast
Extracted proteinSmall amount of pellet
Isolated colonies
Sample Preparation and Measurement Condition - I
22
MALDI TOF mass spectral patterns of TFA extracted proteins of Aspergillus species at growth stage I (A) and growth stage II (B) of sporulation
(A) (B)
0
1000
2000
3000
4000
5000
Inten
s. [a.
u.]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
0.000.25
0.500.75
1.001.25
4x10
Inten
s. [a.
u.]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
0
1000
2000
3000
4000
5000
Inten
s. [a.
u.]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
0
500
1000
1500
2000
Inten
s. [a.
u.]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
0
1000
2000
3000
4000
Inten
s. [a.
u.]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
(a)
(b)
(c)
(d)
(e)
A. fumigatus (BCRC 32149)
A. niger (BCRC 30201)
A. flavus (BCRC 30006)
A. terreus (BCRC 30135)
A. versicolor (BCRC 31488T)
8322
5787
5818
5900
11028
(5942)
7925(8321)
2187
5941 (8335)2836
1565
4146 5773
(11190)93676927
(5798)
(8340)
(1565)
7219
136361116794306967
(1565)
5968
(1565)
8338
m/z
Inte
nsity
(a.
u.)
2000 4000 6000 8000 10000 12000 16000140000
1000200030004000
0
500
10001500
2000
x104 4000 6000 8000 10000 12000 14000 160002000
4000 6000 8000 10000 12000 14000 160002000
4000 6000 8000 10000 12000 14000 160002000
4000 6000 8000 10000 12000 14000 160002000
0.000.25
0.750.50
1.001.25
10002000
0
300040005000
10002000
0
300040005000
m/z
x104
0.0
0.5
1.0
1.54x10
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
0.0
0.5
1.0
1.5
4x10
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
0.00
0.25
0.50
0.75
1.00
1.254x10
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
0.0
0.2
0.4
0.6
0.8
4x10
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
0500
10001500200025003000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
(a)
(b)
(c)
(d)
(e)
A. fumigatus (BCRC 32149) 14226
A. niger (BCRC 3201)
A. flavus (BCRC 30006)
A. terreus (BCRC 30135)
A. versicolor (BCRC 31488T)
12250
2836
5943 11259
10287 11942
13630
4143(4513)
7897
(5944)10835
672214402
7091
(14251)
1565
8232
4423 5974 6816(10545)
(5940)
96103950
11216
m/z4000 6000 8000 10000 12000 14000 16000 18000
2000
02000
50010001500
25003000
x104
x104
x104
4000 6000 8000 10000 12000 14000 16000 180002000
4000 6000 8000 10000 12000 14000 16000 180002000
4000 6000 8000 10000 12000 14000 16000 180002000
4000 6000 8000 10000 12000 14000 16000 180002000
0.000.250.50
1.001.25
0.75
0.0
0.5
1.5
1.0
0.0
0.5
1.5
1.0
0.00.20.40.6
0.8
m/z
2323
MALDI-TOF MS of living and wet-heating inactivated C. albicans
02000400060008000
Inten
s. [a.u
.]2000 4000 6000 8000 10000 12000 14000 m/z
02000400060008000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
0
2000
4000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
0.000.250.500.75
4x10
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
BCRC 20512
0200040006000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
BCRC 21538
02000400060008000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
BCRC 22063
6470
13262119881141210380876485596872
6616
1327587708567
6476 6879
6624 1199710391 11391
6476
6623
6878
1326987708566
1199410390 11413
BCRC 20513
13242
2588
66241010 4175 10391
662645241010
2632BCRC 20519
41766475
5057 5958
29351779
2369
11997877275736474
1751
BCRC 20518
13240
2588
6875662048211008
28514175 6346
2990 33988542
87701139995107449
140521141995125998
3399
951775766880
*** *
* *
*
13745
12848
*
*
*
*
(m/z)
Homogenization with 5% formic acid and 35% ACN
Living intact yeast cells
Inactivated yeast cells
*
*
*
*
*
*
*
*
*
*
*
*
*
*
**
*
*
*
*
*
*
*
*
*
*
*
**
*
*
*
*
*
*
*
*
*
*
*
*
* : Specific reproducible m/z in living and inactivated yeast cells (more than 2 strains)
23
Heat-extracted proteins may be removed by washing
2424
Strain-to strain reproducibility of Candida albicans (Reference strains from BCRC)
02000400060008000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
BCRC 20513
13242
2588
66241010 4175 10391
0
2000
4000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
662645241010
2632BCRC 20519
0.00.51.0
4x10
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
BCRC 20512
1008
2588
6620
41766475
5057 5958
29351779
2369
10391 11994 12845 140588770
853864686880
40682987
3514
11997877275736474
1751
02000400060008000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
BCRC 20518
13240
2588
6875662048211008
28514175 6346
2990 33988542
87701139995107449
140521141995125998
3399
59655048
41783397
2849
951775766880
*
*
*
*
*
** *
* **
*
*
13745
12848
* : Specific reproducible m/z in more than 2 strains of the same Canddia species (small variation range)
* *
*
**
*
*
*
*
**
*
*
*
*
*
*
*
*
*
*
*
*
*
*
*
*
**
*
**
*
*
*
* *
2525
0.0
0.5
1.04x10
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
0
2000
4000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
0
2000
4000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
0.0
0.5
1.04x10
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
CBS 5990
13239
01000200030004000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
CBS 27306880
CBS 6589
CBS 2718
CBS 6431
13240
687625861011 3515 6622
8543 10383
1011
2585
35134065
66186871
2586
6621
68751011
3396
6467
11876 1201310398
6473
6207
35171011 34008574 8774
13251
120031141610394
662464692587
1011 878185653398
3516*
*
*
*
*
*
*
*
*
*
*
**
*
**
*
*
*
**
*
*
*
*
**
*
1344613738
11417
876585644053
11986113994066
64668538 8764 10375 11984 13235
6470 8768
Strain-to strain reproducibility of Candida albicans (Reference strains from CBS)
*
*
*
*
*
*
*
*
*
*
*
*
*
*
*
*
* **
*
*
*
*
*
*
* : reproducible m/z
2626
Strain-to-strain reproducibility of different Candida speciesCandida tropicalis
Candida glabrata
BCRC 20560
0100020003000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
BCRC 21436
7213
69201011
28433564
30182495
4503
0200040006000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
1011 72113019 3492
44982492
020004000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
CBS 2175
10116589
69338868
9606
10778
2972
3314
4167 5392 7666 1092611095
2148
6221
5843
0200040006000
Inten
s. [a.u
.]
2000 4000 6000 8000 10000 12000 14000 m/z
10777
9607
886476626929
65855389
48181011
2147 2970 41645839
6218
3310
10922
11101
CBS 860
High strain-to-strain reproducibility
*
*
*
*
*
*
*
*
*
**
*
**
* *
*
**
**
*
*
**
*
*
* *
*
*
*
*
*
*
* *
*
* : reproducible m/z
7571
85698812
27270 50 100 150 200
0
2
4
6
8
10
12
14
16
18
20
Variance Weighted Distance Between Cluster Centers
Candida tropicalis_BCRC-21436
Candida tropicalis_BCRC-20560
Candida albicans_BCRC-20512
Candida albicans_CBS-2718
Candida albicans_BCRC-20518
Candida albicans_CBS-5990
Candida albicans_CBS-6589
Candida albicans_BCRC-20513
Candida albicans_CBS-2730
Candida albicans_CBS-6431
Candida albicans_BCRC-20519
Candida krusei_BCRC-21321
Candida krusei_BCRC-21720
Candida glabrata_CBS-860
Candida glabrata_CBS-2175
Candida lusitaniae_BCRC-20326
Candida lusitaniae_CBS-7270
Candida parapsilosis_BCRC-21253
Candida parapsilosis_BCRC-21544
Dendrogram of Data with Preprocessing: 2nd Derivative (order: 2, window: 15 pt) + Normalize (1-Norm, Area = 1) + Autoscale
Cluster analysis of mass spectral patterns of different strains of Candida species
19 tested samples 27
Discrimination to the species level attributed to small variations of both qualitative m/z values and quantitative relative intensities
ʳ * tear tube 32x after 1d-L12-1800shots-200kDa\0_L12\1\1SLin, "Baseline subt."
0
1
25x10
Inte
ns. [
a.u.
]
ʳ
* tear from paper 4x 1800 shots\0_I20\1\1SLin, "Baseline subt."
0.0
0.5
1.05x10
Inte
ns. [
a.u.
]
ʳ
* Jung Hung Wang N20-OS-1800a\0_N20\1\1SLin, "Baseline subt."
0
1
4x10
Inte
ns. [
a.u.
]
ʳ
* Jung Hung Wang N21-OD-1800a\0_N21\1\1SLin, "Baseline subt."
0.0
0.5
1.0
4x10
Inte
ns. [
a.u.
]
2000 4000 6000 8000 10000 12000 14000 16000m/z
ʳ * tear 1x with SA-std protocol-two layer dried 52 per power-3300 adding\0_A8\1\1SLin, "Basel...
0
1
2
3
4
4x10
Inte
ns. [
a.u.
]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
Artificial tears for simulation and human tears/contact lens solution
Artificial ears for simulationHuman tear/contact lens solution from
healthy volunteer
Section C(II)_Paper ID : 230 Applications to food industry, environmental monitoring, and healthcare
Left eye
Right eye
Schirmer test strips
Glass capillary tube
Contact lens solution
Contact lens solution
14296
7148
Lysozyme (chicken egg) + 1 H+
Lysozyme + 2 H+
Human lysozyme + 1 H+
14696
Tears and contact lens solution from patients with corneal ulcer
ʳ * corneal ulcer sample 4x-C3-1800\0_C3\1\1SLin, "Baseline subt."
0.0
0.5
1.0
1.55x10
Inte
ns. [
a.u.
]
ʳ
* 6314968-OD-T82-1800a-16x\0_O5\1\1SLin, "Baseline subt."
0
2
4
64x10
Inte
ns. [
a.u.
]
ʳ
* 60564348-OS-T81-1800a-2x\0_P20\2\1SLin, "Baseline subt."
0
2
4
6
84x10
Inte
ns. [
a.u.
]
2000 4000 6000 8000 10000 12000 14000 16000 18000m/z
Tears collected by glass capillary tube Immersion liquids of non-washed worn contact lenses (without proteolytic enzyme)
Section C(II)_Paper ID : 230 Applications to food industry, environmental monitoring, and healthcare
ʳ * C17-CL 002-left-2ul\0_C17\1\1SLin, "Baseline subt."
0.0
0.5
1.0
4x10
Intens. [a.u.]
ʳ
* C18-CLB 002-right-2ul\0_C18\1\1SLin, "Baseline subt."
0.00.5
1.0
1.5
4x10
Intens. [a.u.]
ʳ
* G14-CLB 001-left 2ul\0_G14\1\1SLin, "Baseline subt."
0.0
0.5
5x10
Intens. [a.u.]
ʳ
* E15-CLB 001-right 2ul-after cali\0_E15\1\1SLin, "Baseline subt."
0
2
4
4x10
Intens. [a.u.]
4000 6000 8000 10000 12000 14000 16000 18000m/z
Left eye
Right eye
Left eye
Right eye
Patient A
Patient B
Patient C
Patient D
Patient D
Patient E
Patient E
Infected eye
Infected eye
Infected eye
14678
7339
Human lysozyme (1 H+)
Human lysozyme (2 H+)
14695
7339
Human lysozyme (1 H+)
Human lysozyme (2 H+)
Abnormal marker*
*
*
*
*
*
*
14695
14694
14680
ʳ * A16-1800a\0_A16\1\1SLin, "Baseline subt."
0
2
4
6
4x10
Intens. [a.u.]
ʳ
* C18-1800a\0_C18\1\1SLin, "Baseline subt."
0
2
4
6
84x10
Intens. [a.u.]
ʳ
* 7-P aeruginosa 10944-blood culture\0_B7\1\1SLin, "Baseline subt."
0
1
2
3
4x10Intens. [a.u.]
5000 10000 15000 20000 25000 30000 35000m/z
ʳ * 4-S aureus 14957-TSA\0_B4\1\1SLin, "Baseline subt."
0.0
0.5
1.0
4x10
Intens. [a.u.]
ʳ
* 5-S aureus 14957-chocolate\0_B5\1\1SLin, "Baseline subt."
0.0
0.5
1.0
5x10
Intens. [a.u.]
ʳ
* 6-S aureus 14957-blood culture\0_B6\1\1SLin, "Baseline subt."
0
2
4
6
4x10
Intens. [a.u.]
4000 6000 8000 10000 12000 14000 16000 18000 20000m/z
Common pathogens of patients with infectious corneal ulcer
Section C(II)_Paper ID : 230 Applications to food industry, environmental monitoring, and healthcare
P. aeruginosaS. aureus TSA culture TSA culture
Blood agar culture
Chocolate agar culture Chocolate agar culture
Blood agar culture
Early stage of cultivation (1-3 days) at 35 oC
9093*7582*
13956*
6887*
3882*9631*2311*
6887*
6886*
9634*
9626*
3881*
3876*
9092*
9093*
13956*
13955*
7581*
7582*
*Identification marker
SwissProt database search by Mascot software
31