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A. Phos-tag™ - Phosphorylated Protein Analysis
1.Phos-tag™ Acrylamide
2.SuperSep™ Phos-tag™
3.Phos-tag™ Biotin
4. Phos-tag™ Mass Analytical Kit
5. Phos-tag™ Agarose
B. Protein Electrophoresis
1. SuperSep™ - Precast Polyacrylamide Gel
2. Easy Separator™
3. Reagents for Gel Preparation
4. Protein Size Markers
5. Premixed Buffers - Running Buffers & Sample Buffers
6. Staining Reagents
a. Negative Gel Stain MS Kit
b. Silver Staining Kits
c. CBB R-250 Stain Kits
C. Western Blotting
1. Blocking Reagents
2. Wash Solutions
3. Labeled 2nd Antibodies
4. Multi Capture HRP - Antibody Detection Reagent
5. Immuno-enhancer - Accelerator of Antigen-Antibody Reactions
6. Stripping Reagent
7. Coloring Reagents
8. ImmunoStar LD Chemiluminescence Reagent
D. MS Analysis
1. Reagents for Sample Pretreatment
2. Matrix for MALDI-TOFMS Analysis
3. In-gel Digestion Enzymes
4. LC/MS Solvents
5. MALDI-MS Calibrants










A. Phos-tag™ - Phosphorylated Protein Analysis
     Phos-tag™ Series
Basic structure of Phos-tag™
Zinc ion or Manganese ion

Phos-tag™ is...
Phos-tag™ is a functional molecule that binds specifi cally phosphorylated ions.

Features
Selectivity of binding of a phosphate ion (2-) is much higher than that of other anions.
A stable complex is formed under physiological conditions (pH 5 to 8).
∗: In case of Zn2+ is used as M2+.

Description Application
Phos-tag™ Acrylamide Specifi c separation of phosphorylated proteins corresponding to the degree of phosphorylation
Phos-tag™ Biotin Specifi c detection without any anti-phosphorylated antibodies on western blot
Phos-tag™ Agarose Enrichment, separation and purifi cation of phorphorylated proteins using column chromatography
Phos-tag™ Mass Analytical Kit Analysis: Applicable to MALDI-TOF/Mass analysis of phosphorylated compounds in positive mode using an isotopic Zn2+-Phos-tag™ complex
Phos-tag™ was developed by Department of Functional Molecular Science at Hiroshima University.

1. Phos-tag™ Acrylamide   (Please see the product list on the page #5.)

Separation of phosphorylated protein isoforms using SDS-PAGE
Phos-tag™ Acrylamide is added when a resolving gel of SDS-PAGE is prepared. Since Phos-tag™ in the gel can trap phosphorylated proteins and the migration speed of phosphorylated proteins decreases, they can be separated from non-phosphorylated proteins. (Phos-tag™ SDS-PAGE).

Features Principle
Recognition of all
phosphorylated forms of Ser /
Thr / Tyr
Applicable to Western blotting
and Mass analysis
Simultaneous detection of
phosphorylated- and nonphosphorylated
protein using
the total antibody without the
anti phosphorylated protein.
Application
Time course of phosphorylation by using the Tyrosine Kinase Abl
Phosphorylated tyrosine was prepared by GST binding protein of tyrosine kinase Abl and the substrate peptide (Abltide)
and separated with conventional SDS-PAGE and Phos-tag™ SDS-PAGE, respectively.
 

2. SuperSep™ Phos-tag™ - Precast Phos-tag™ acrylamide Gel

This is a precast gel added with Phos-tag™ Acrylamide in advance. It
can be used immediately after opening the package. It contains zinc
as a supplemental metal. It has excellent storage stability by its neutral
gel buffer. Sharp bands can be obtained.

Features
Phosphorylated- and non-phosphorylated proteins are efficiently
    separable with sharp bands.
Long-term stability (Stable for 6 months)

Physical Properties
Plate size 100 x 100 x 3 (mm)
Gel size 90 x 85 x 1 (mm)
Well number 13
Well volume (μL) 30

Application 1
[Buffer] Tris-Glycine-SDS electrophoresis buffer
[Sample] 1 : α-casein / 2 : Dephosphorylated α-casein
[Condition] Constant current 40 mA for 35 min.
[Staining] Quick CBB Staining
[Destaining]
  Destaining with methanol-acetic acid decoloring agent for 30 min.
  α-casein was processed with alkaline phosphatase and subjected to electophoresis
  using SuperSep™
  Phos-tag™ and 12.5% SuperSep™ as a control. Band shift by dephosphorylation
  was observed.

Application 2
[Buffer]
  Tris-Glycine-SDS electrophoresis buffer
[Sample]
  M: WIDE-VIEW™ Prestained Protein Size Marker III*.
  1:   0 min. β-casein (AP treatment)
  2: 15 min. β-casein (AP treatment)
  3: 30 min. β-casein (AP treatment)
  4: 45 min. β-casein (AP treatment)
  5: 60 min. β-casein (AP treatment)
[Condition] Constant current 35 mA for 60 min.
[Staining] Quick-CBB Staining
[Destaining]
  Deionized water with microwave
  β-casein was dephosphorylated over time.
  Bands of dephosphorylated β-casein increased over time.
Application Description Wako Cat. No. Package Size
Separation of phosphorylated and
non-phosphorylated protein by SDS-PAGE
SuperSep Phos-tag™ (50 μmol/L)
12.5 %, 13 well
5 sheets 195-16391
∗: Note
Prestain marker does not refl ect the molecular weight, when a gel containing Phos-tag™ is used. Please use the result as an index of transfer efficiency at Western blotting. WIDE-VIEW™ Prestained Protein Size Marker III (See the page #8.) is used in this marker.
At Western blotting, Zn2+ must be sufficiently removed with transfer buffer containing 10 mM EDTA. Please wash out the gel 10 min x 3 times. Subsequently, perform the same procedure 10 min. X 1 using transfer buffer without EDTA, then transfer the gel.

3. Phos-tag™ Biotin
Detection of phosphoprotein for Western blotting

This is biotin-bound Phos-tag™ used for detection of phosphoprotein by
Western blotting.
Features
All phosphoprotein can be detected.
Procedures of experiment are similar to those in ordinary Western blotting.
It can be conveniently used even when target anti-phosphorylated Thr/
Ser antibody is not available !
BTL-104 and BTL-105 have linkers with different length, but their usage are similar.
BTL-104 is recommendable as the fi rst choice because of its high solubility.

4. Phos-tag™ Mass Analytical Kit
Detection sensitivity of MALDI-TOF/Mass improved Kit Contents
Before use, Phos-tag™ Mass Analytical Kit is mixed with samples for
MALDI-TOF/Mass analysis. Phosphorylated molecule-Phos-tag™
complex is detected in a positive mode, and phosphorylated molecule
usually difficult to detect can be detected with improved sensitivity.
Phos-tag™ MS-101L (5 mg x 1 vial)
    ([C27H29N6O64Zn2]3+ MW : 581.4)

Phos-tag™ MS-101H (5 mg x 1 vial)
    ([C27H29N6O68Zn2]3+ MW : 589.4)

Phos-tag™ MS-101N (10 mg x 1 vial)
    (C27H29N6OZn2]3+ MW : 584.3)

Features
Detection sensitivity of phosphorylated molecule is improved.
101L, 101H and 101N have different types of Zinc.
101N contains many isotopes because it uses natural Zinc that has complex mass spectrum. It is suited for evaluation of condition.
101L and 101H contain Zinc with mass numbers of 64 and 68, respectively. They are suited for detection of phosphate group.

5. Phos-tag™ Agarose
Purification of phosphorylated protein by affinity chromatography
Fill Phos-tag™ Agarose in a column for use. It can be used for separation, purification and concentration of phosphorylated proteins. Because it is free from a surfactant or reducing agents, phosphorylated protein can be obtained in a condition similar to in vivo one.
Features
Phosphorylated proteins can be purified in 1 hour.
The proteins can be trapped in physiological
condition (pH 7.5).
No reducing agent or surfactant is necessary.
Application Description Manufacturer's
product No.
Wako Cat. No. Package
Size
Separation of phosphorylated /
nonphosphorylated proteins by SDS-PAGE
Phos-tag™ Acrylamide (Nard Institute) AAL-107M 300-93523 2 mg
AAL-107 304-93521 10 mg

Phos-tag. Acrylamide 5 mM Aqueous Soln.(Nard Institute)
AAL-107S1 304-93526 0.3 mL
They can be used in place of anti-
phosphorylated antibody.
Any amino acids can be detected.
Phos-tag™ Biotin BTL-104 (Nard Institute) BTL-104 301-93531 10 mg
Phos-tag™ Biotin BTL-105 (Nard Institute) BTL-105 308-93541 10 mg
Phos-tag™ Biotin BTL-111S1
(Nard Institute)
BTL-111S1 308-97201 0.1 mL
Analysis by MALDI-TOF/Mass Phos-tag. Mass Analytical Kit (Nard Institute) MS-101KIT 305-93551 1 set
Purification of phosphorylated proteins Phos-tag™ Agarose (Manac) AG-501 302-93561 0.5 mL
AG-503 308-93563 3 mL

B. Protein Electrophoresis

1. SuperSep™ - Precast Polyacrylamide Gel
SuperSep™ is a precast polyacrylamide gel for electrophoresis of proteins or nucleic acids. Because the gel does not
contain SDS, it can be used for SDS-PAGE in SDS-containing buffer, and for Native-PAGE in buffers that do not contain
SDS. 12 kinds of precast gels are available.
Features
Safety
High separation efficiency
Superior stability (Stable for 6 or 9 months, depending on item)
Product Specificationz
Well 13 17
Well volume (μL) 30 25
Mass of Total Protein (μg)∗ 3.3 - 6.5 1.3 - 3.9
Plate Size 100 x 100 x 3 (mm)
Gel Size 90 x 85 x 1 (mm)

∗Target amount of protein that can be separated efficiently with SuperSep™ gel
Note:
In order to obtain sharp bands, apply a sample as little as possible slowly to the bottom of each well. When much sample is applied, resultant bands may be
blurred. It is advisable to rinse inside of each well with a syringe or a fi ne pipette, before applying samples.
Target of separation
Please select appropriate gel concentration, with reference to the following separation pattern.
References:
Kosaka, N., et al. : J. Biol, Chem. 285, 17442 (2010)
Prabhala, RH., et al. : Blood, 115, 5385 (2010)
Shimada, Y., et al. : Appl. Envir. Microbiol., 76, 5892 (2010)
Gopinaht, SC., et al. : Mol. Cancer Res., 8, 1536 (2010)
Hayashi-Tanaka, Y., et al. : Nucleic Acids Res., 39, 6475 (2011)
Koumura, A., et al. : J. Pharmacol. Exp. Ther., 338, 337 (2011)
Taguchi, K., et al. : J. Cell Biol., 194, 643 (2011)

Description well Package Size Wako Cat. No. Shelf life
SuperSep™ Ace, 6% 13 10 sheets 195-15171 6 months
17 10 sheets 192-15181
SuperSep™ Ace, 7.5% 13 10 sheets 198-14941 9 months
17 10 sheets 191-14931
SuperSep™ Ace, 10% 13 10 sheets 195-14951
17 10 sheets 192-14961
SuperSep™ Ace, 12.5% 13 10 sheets 199-14971
17 10 sheets 196-14981
SuperSep™ Ace, 15% 13 10 sheets 193-14991
17 10 sheets 190-15001
SuperSep™ Ace, 5-12% 13 10 sheets 199-15191 6 months
17 10 sheets 192-15201
SuperSep™ Ace, 5-20% 13 10 sheets 197-15011 9 months
17 10 sheets 194-15021
SuperSep™ Ace, 10-20% 13 10 sheets 191-15031
17 10 sheets 198-15041
SuperSep™ Ace, 15-20% (Tricine Gel) 13 10 sheets 198-15301 6 months
SuperSep™, 12.5%, 2D 2D 10 sheets 190-13301 9 months
SuperSep™, 5-20%, 2D 2D 10 sheets 197-13291
SuperSep™, 10-20%, 2D 2D 10 sheets 192-14721
∗Shelf life after manufacturing date

2. Easy Separator™
This is an electrophoresis tank for SuperSep™ precast
polyacrylamide gel. Attachment and removal of the gel are
easy.
Turn the knob after electrophoresis, then running buffer filled
inside in the tank easily falls down. So, the gel can be
removed directly.
Features
Easy set-up of SuperSep™ by turning the knob.
Easy removal of SuperSep™ without disposing the inside
running buffer after electrophoresis.
Electrophoresis can be performed with a minimum amount of
running buffer (250 - 350 mL)
Description Wako Cat. No. Package Size
Easy Separator™ 058-07681 1 set

3. Reagents for Gel Preparation
Followings are reagents used to prepare polyacrylamide gel for protein electrophoresis. Various kinds of reagents such as
powder and ready-to-use solution are available.

Description <Grade∗> Wako Cat. No. (Package Size)
Acrylamide, 99.0+% (cGC) <for Electrophoresis> 017-08012 (25 g) ; 019-08011 (100 g) ; 011-08015 (500 g)
Acrylamide-HG, 99.9+% (cGC) <for Electrophoresis> 013-10023 (10 g) ; 017-10021 (100 g) ; 019-10025 (500 g)
Acrylamide-HG, 99.9+% (cGC) <for Molecular Biology> 013-20751 (100 g) ; 015-20755 (500 g)
30 w/v% Acrylamide Solution, 37.5 : 1 <for Electrophoresis> 016-15915 (500 mL)
30 w/v% Acrylamide Solution-HG, 37.5 : 1 <for Proteomics> 010-22645 (500 mL)
30 w/v% Acrylamide Solution, 29 : 1 <for Electrophoresis> 014-21705 (500 mL)
Separating Gel Buffer Solution ( x 4)
(0.4 w/v% SDS in 1.5mol/L Tris-HCl, pH 8.8)
<for Electrophoresis>
192-11041 (250 mL)
Stacking Gel Buffer Solution ( x 4),
(0.4 w/v% SDS in 1.5mol/L Tris-HCl, pH 6.8)
<for Electrophoresis>
199-11051 (250 mL)
N,N'-Methylenebis (acrylamide) -HG,
99.0+% (Titration)
<for Electrophoresis>
132-08171 (1 g) ; 130-08172 (25 g) ; 138-08173 (100 g)
N,N'-Methylenebis (acrylamide) -HG,
99.0+% (Titration)
<for Molecular Biology>
132-15082 (25 g) ; 134-15081 (100 g)
N,N,N',N'-Tetramethylethylenediamine
(TEMED), 99.0+% (cGC)
<for Electrophoresis>
205-06313 (25 mL)
Ammonium Peroxodisulfate (APS),
99+ % (Titration)
<for Electrophoresis>
012-08023 (10 g) ; 016-08021 (100 g)
10 w/v% Ammonium Peroxodisulfate Soln.
(10 w/v% APS)
<for Electrophoresis>
019-15922 (25 mL)
Ribofl avin, 98.0+ % (Absorptiometry) <for Electrophoresis> 181-00581 (1 g)
2-Amino-2-hydroxymethyl-1,3-propanediol
(Tris), 99.0+ % (Titration)
<for Molecular Biology>
019-20091 (100 g) ; 011-20095 (500 g) ; 015-20093 (1 kg)
2-Amino-2-hydroxymethyl-1,3-propanediol 999,
99.9+ % (Titration)
<for Biochemistry>
015-16384 (100 g) ; 013-16385 (500 g) ; 011-16381 (1 kg) ; 017-16383 (5 kg)
Sodium Dodecyl Sulfate (SDS),
95.0+ % (GC) ; 98.0+ % (Titration)
<for Biochemistry>
197-07142 (25 g) ; 199-07141 (100 g) ; 191-07145 (500 g)
Sodium Dodecyl Sulfate (SDS), 95.0+ % <for Molecular Biology> 190-13982 (25 g) ; 192-13981 (100 g) ; 194-13985 (500 g)
10% SDS Solution <Nippon Gene> 311-90271 (100 mL) ; 313-90275 (500 mL)
* : for Molecular Biology - A series of reagents with DNase and RNase activities checked.

4. Protein Size Markers
No. Description Wako Cat. No. Package Size

1
WIDE-VIEW™ Prestained Protein Size Marker III (11 - 245 kDa) 230-02461 500 μL (for 100 tests)
2 WIDE-VIEW™ Prestained Protein Size Marker (15- 140 kDa) 230-02221 500 μL (for 50 - 100 tests)
3 Molecular Weight Marker, Low Range (3.5 - 42 kDa) 294-63101 1 mL (for 200 tests)
4 Molecular Weight Marker, Middle Range (14 - 79 kDa) 131-14511 1 mL (for 200 tests)
5 Molecular Weight Marker, Wide Range (6.5 - 180 kDa) 296-63301 1 mL (for 200 tests)

5. Premixed Buffers
This is a series of ready-to-use
premixed buffers for protein
electrophoresis. They can be used
immediately after dilution.

     Running Buffers for SDS-PAGE
Description Application Wako Cat. No. Package
Size
Composition
Running Buffer Solution ( x 10) (Tris / Glycine / SDS Buffer) Laemmli method Tris-Glycine
Running Buffer containing SDS
184-01291 1 L
Composition: 250 mmol/L Tris, 1,920 mmol/L Glycine, 1.0% (w/v) SDS
SDS-PAGE 10 x Running Buffer <Nippon Gene> Laemmli method Tris-Glycine
Running Buffer containing SDS
312-90321 1 L
Composition: 250 mmol/L Tris, 1,920 mmol/l Glycine, 1.0% (w/v) SDS 318-90323 5 L
10 x Tris-Glycine Buffer Native-PAGE, Transfer Buffer 201-18601 1 L
Composition: 0.25 mol/L Tris 1.92 mol/L Glycin
Tricine Running Buffer Solution ( x 10) (Tris / Tricine / SDS Buffer) Running Buffer for Tris-Tricine gel 200-17071 1 L
Composition: 0.5 M Tris / 0.5 M Tricine /1% SDS
     Electrophoresis Sample Buffers
Laemmli Sample Buffers -Three types are available: 1 new buffers added with non-hazardous chemical, 3-mercapto-1,2-
propanediol; 2 buffers with 2-mercaptoethanol (2-ME+); 3 buffers without 2-mercaptoethanol (2-ME-).
Description Application Wako Cat. No. Package
Size
Composition
Sample Buffer Solution (2-ME+) ( x 2) Laemmli Sample Buffer with
2-Mercaptoethanol
196-11022 25 mL
Composition: 0.125 mol/L Tris-HCl (pH 6.8), 4w/v% SDS, 20 w/v% Glycerol, 196-11022 25 mL
0.002 w/v% BPB, 10 vol% 2-Mercaptoethanol
Sample Buffer (2-ME+) ( x 4) 191-13272 25 mL
Composition: 0.25 mol/L Tris-HCl (pH 6.8), 8 w/v% SDS, 40 w/v% Glycerol, 191-13272 25 mL
0.02 w/v% BPB, 20 vol% 2-Mercaptoethanol
25 mL
Sample Buffer (2-ME-) ( x 2) Laemmli Sample Buffer without
2-Mercaptoethanol
193-11032 25 mL
Composition: 0.125 mol/L Tris-HCl (pH 6.8), 4w/v% SDS, 20 w/v% Glycerol, 193-11032 25 mL
0.002 w/v% BPB
Sample Buffer (2-ME-) ( x 4) 198-13282 25 mL
Composition: 0.25 mol/L Tris-HCl (pH 6.8), 8 w/v% SDS, 40 w/v% Glycerol, 198-13282 25 mL
0.02 w/v% BPB
Sample Buffer with 3-Mercapto-1,2-propandiol ( x 2) Laemmli Sample Buffer
containing 3-Mercapto-1,2-
propanediol (non-hazardous
chemical) as substitute for 2-ME
199-16132 25 mL
Composition: 0.125 mol/L Tris-HCl (pH6.8), 4 w/v% SDS, 20 w/v% Glycerol, 199-16132 25 mL
0.01 w/v% BPB, 10 v/v% 3-mercapto-1,2-propandiol
Sample Buffer with 3-Mercapto-1,2-propanediol ( x 4) 196-16142 25 mL
Composition: 0.25 mol/L Tris-HCl (pH 6.8), 8 w/v% SDS, 40 w/v% Glycerol, 196-16142 25 mL
0.02 w/v% BPB, 20 v/v% 3-mercapto-1,2-propandiol

6. Staining Reagents
     Negative Gel Stain MS Kit
It is known that protein bands separated by SDS/polyacrylamide gel electrophoresis (SDS-PAGE) can be visualized as
transparent bands against the background of milky white gel stained by negative gel stain containing a Zn/imidazol reagent.
We have improved the method using a new imidazol derivative reagent, which allows a clear and stable image of protein
bands on the gel as sensitive as that by silver staining, in as little as 10 minutes. The staining technique is useful to obtain
the clear and sensitive resolution pattern of the gel before immunoblotting as well as to excise and purify the band of
interest from the gel without signifi cant deterioration of amino acid residues for the subsequent studies of protein such as
sequencing and mass analysis of peptide.

Staining Principle MALDI-TOF/MS of rat phosphorylase
Staining uses insoluble opaque ZnIm2 generated by reaction
of free Zn2+ and imidazole.
Protein and protein-SDS complex react with Zn2+. The moiety
reacted with Zn2+ remains clear because insoluble ZnIm2
does not deposit on it, but the moiety other than protein
becomes opaque because ZnIm2 deposits on it. This type of
staining is called negative staining, because unlike other
staining, background is stained and protein is not stained.
Features Kit Contents
High sensitive (3 ng) and Quick detection (10 minutes).
Applicable to the subsequent studies of protein such as sequencing and
mass analysis of peptide.
Repeatable staining and destaining are acceptable.
After destaining, the gel is applicable to Western blotting.
Staining Solution A 500 mL x 1 bottle
Staining Solution B 500 mL x 1 bottle
De-staining Solution 500 mL x 1 bottle
Description Grade Wako Cat. No. Package Size
Negative Gel Stain MS Kit for Electrophoresis 293-57701 20 tests

     b. Silver Staining Kits
3 types of high sensitive Silver Staining Kits are available.
Principle
Silver ion selectively binds to sulfhydryl group (-SH), amino group (-NH2), hydroxyl group (-OH), or carboxyl group (-COOH),
which consist of protein. Especially, reaction with sulfhydryl group (-SH) occurs preferentially. Silver ion forming complex with
ammonium ion in the reagent binds to protein, via substitution of ammonium ion and sulfhydryl group in protein.

Silver Stain MS Kit
This kit is produced for mass spectrometric sequencing of proteins separated by polyacrylamide gel electrophoresis, based
on the method described by Shevchenko, et al. Due to the omission of glutaraldehyde treatment, proteins are rarely
modifi ed chemically and therefore can be detected at a sub-nanogram level on the electrophoretic gel.
Kit Contents
Enhancing Stock Soln. 200 mL x 1
Staining Stock Soln. 200 mL x 1
Developing Stock Soln. 100 mL x 1
Developing Powder 20 g x 1
Stopper 200 mL x 1
De-staining Soln. A 50 mL x 1
De-staining Soln. B 50 mL x 1
Application
MALDI-TOF/MS of rabbit phosphorylase
The band was excised and treated with
Lysyl Endopeptidase® (#125-02543).
Following the in-gel digestion and
preparation, the sample was analyzed on
MALDI-TOF mass spectrometer. (These
data were provided by Dr. Y. Wada at
Osaka Medical Center, Japan.)
Grade Description Package Size Wako Cat. No.
for Electrophoresis Silver Stain MS Kit 20 tests 299-58901

Silver Stain Kit Wako
Silver Staining Kit Wako has high sensitivity and reproducibility.
Staining fi nishes in 100 minutes.
Kit Contents
Fixing Stock Soln. (Glutaraldehyde) 200 mL x 2
Enhancing Stock Soln (Dithiothreitol) 10 mL x 1
Staining Soln. A (Silver nitrate) 100 mL x 1
Staining Soln. B (Ammonia & Sodium Hydroxide) 100 mL x 1
Developing Stock Soln. 100 mL x 1
(Formaldehyde & Citric Acid)
Grade Description Package Size Wako Cat. No.
for Electrophoresis Silver Stain Kit Wako for 10 sheets 299-13841

Silver Stain II Kit Wako
An improved type Silver Staining kit which can stain
protein bands in 1 hour. It contains Stopper, which can be
used for adjustment of staining for your intended use.
Kit Contents (100 mL each x 1 bottle)
Fixing Stock Soln. (Methanol & Acetic Acid)
Enhancing Stock Soln. (Dithiothreitol & Glutaraldehyde)
Staining Soln. A (Silver Nitrate)
Staining Soln. B (Ammonia & Sodium Hydroxide)
Developing Stock Soln. (Formaldehyde & Citric Acid)
Stopper (Citric Acid)
Grade Description Package Size Wako Cat. No.
for Electrophoresis Silver Stain II Kit Wako for 10 sheets 291-50301

Wako's Silver Staining Kits
(Note 1) Since these kits do not contain this reagent, please prepare it by yourself. When you use Negative Gel Stain MS Kit, it is not necessary to fi x samples.
However, higher sensitivity will be obtained with 50 % ethanol fixing for 5 minutes
(Note 2) Brown colored staining parts are excised in case of silver staining. On the other hand, black colored parts against a black paper are excised in case of
negative gel staining.
(Note 3) When used in subsequent MS analysis, de-staining is unnecessary. When used is subsequent Western Blotting, the gel is de-stained.
(Note 4) Lysyl Endopeptidase, Mass Spectrometry Grade (Wako Cat. No. 125-05061; 20 μg x 5) / Trypsin, from Porcine Pancreas, Mass Spectrometry Grade
(Wako Cat. #202-15951; 20 μg x 5) are available from Wako. (Please see the page #22.)
(Note 5) Each Silver Stain Kit does not contain deionized water, methanol and acetic acid. Please prepare by yourself.
(The above mentioned reagents are not necessary when you use Negative Gel Stain MS Kit.)

     c. CBB R-250 Stain Kits
Quick CBB
Quick CBB is an upgraded version of the conventional CBB stain
using CBB R-250. After fixing treatment, proteins are stained by
immersing the gel in Quick CBB solution. Protein bands can be
detected in a short time after SDS-PAGE treatment because no
destaining procedure with acetic acid is necessary.

Features
Detectable in 50 minutes.
Easy preparation.
(Prepare Staining Soln. by mixing Soln. A and B at a ratio of 1 : 1)
10 minute detection with microwave. (Application 2)
Grade Description Package Size Wako Cat. No.

for Electrophoresis

Quick CBB

  <Kit Contents>
   1. Staining Solution A (1 L x 1)
   2. Staining Solution B (1 L x 1)


2 L

299-50101

Quick-CBB PLUS
This is a protein staining kit which is a further upgraded version of Quick-CBB. In comparison with conventional Quick-CBB,
fi xing procedure is not required and organic solvents such as methanol and acetic acid are not used. The mixing procedure
is unnecessary as all the required solutions for staining are contained in a single bottle. There are other improvements,
such as the removal of background coloration. As in conventional Quick-CBB, destaining procedure is not required.
Features
No fixing procedure and no organic solvents are necessary.
A single-bottled ready-to-use protein staining kit
Simple and quick staining without the coloration on background.

Application 1
Regular Protocol
Wash: Deionized water 100 mL
↓ (5 min. x 3 times)
Staining: Quick-CBB PLUS 25 mL
↓ (30 - 60 min.)
Wash: Deionized water 100 mL
Grade Description Package Size Wako Cat. No.
for Electrophoresis Quick-CBB PLUS 250 mL 174-00553
1 L 178-00551

Application 2
10 minute protocol by microwave
With Quick-CBB PLUS, staining and decoloring can be completed in about
10 min using microwave. It is very useful when users want the result at once.
Regular Protocol
[Protocol]
1. SDS-PAGE
2. Staining by microwave: for 1 min. x 2 ~ 4 times (In case of Quick-CBB, for 1 min x 1 time)
Pour Quick-CBB PLUS, cover with plastic wrap and heat at 500 W for 1 minute.
Repeat microwaving till the whole gel get colored.
3. Destaining by microwave: 1 min. x 4 ~ 6 times
Discard Quick-CBB PLUS from the tray and pour 100 mL of deionized water to
immerse the gel. Add a balled-up Kimwipe in the tray and heat for a few minutes.
Repeat this destaining procedure with new deionized water.∗
∗ In case of bumping, there is no problem if the gel is immersed in the solution. Leave the gel
in deionized water overnight to lower the background.
(Caution) When the tray is completely covered with plastic wrap, make several holes on the
wrap with needle. The tray becomes very hot. Use gloves and pay full attention
when removing the tray.

C. Western Blotting

1. Blocking Reagents
Blocking reagents used to prevent non-specific absorption of antibody to the transfer membrane. Generally, Bovine Serum
Alubumin (BSA) or Skim Milk are used as a blocking reagent. Skim Milk is a potent blocking reagent, but it is not suitable
for phosphorylated antibodies because they contain a lot of phosphorylated proteins.

Skim Blocker
Skim Blocker is a blocking reagent that prevents non-specifi c absorption to the membrane or ELISA plates. This is a readyto-
use reagent that can be used immediately. Blocking fi nishes by shaking for 30 min. at room temperature.
Assay Procedure
1. After electrophoresis, transfer to a membrane.
2. Wash the membrane with TBS-T, and shake it with Skim Blocker (at room temperature for 30 min.)
3. Wash the membrane with TBS-T and make it react with labeled antibody (at room temperature for 40 min.)
4. Wash the membrane and take photos. (Exposure time: 5 seconds)
Grade Description Package Size Wako Cat. No.
for Blotting Skim Blocker 500 mL 195-16455
Skim Milk Powder 500 g 190-12865
for Biochemistry Gelatin, from Bovine Bone 100 g 074-02761
500 g 076-02765
Albumin, from Bovine Serum, Globulin Free 10 g 019-15101
50 g 015-15103
100 g 013-15104
: Not available for sale in the US.

2. Wash Solutions
Wash solution is a buffer for washing membranes for Western blotting or ELISA plates. It can be easily
prepared by diluting the following solution with distilled water.


Grade Description Package Size Wako Cat. No.
Composition
for Blotting PBS-T, pH7.4 (x10) 1 L 163-24361
Composition: 1,370 mmol/L NaCl, 81 mmol/L Na2HPO4,
27 mmol/L KCl, 15 mmol/L KH2PO4, 1 (w/v)% Tween 20
for Blotting TBS-T, pH7.4 (x10) 1 L 207-18061
Composition: 1,370 mmol/L NaCl, 27 mmol/L KCl,
250 mmol/L Tris, 1 (w/v)% Tween 20
Nippon Gene 20x TBS (pH7.4) 500 mL 317-90371
Composition: 400 mM Tris-HCl (pH 7.4), 3 M NaCl
Nippon Gene 10x TBS (pH7.4) 500 mL 317-90175
Composition: 1,370 mmol/L NaCl, 26.8 mmol/L KCl,
250 mmol/L Tris
Nippon Gene 10xPBS (-) (pH7.4) 500 mL 314-90185
Composition: 1,370 mmol/L NaCl, 81 mmol/L Na2HPO4,
26.8 mmol/L KCl, 14.7 mmol/L KH2PO4/TD>

3. Labeled 2nd Antibodies
Various types of labeled secondary antibodies are available. Anti-mouse IgG, HRP labeled antibody, anti-rabbit IgG and
HRP labeled antibodies are now available.
Application 1 Application 2
Sample: mixture of FLAG-BAP and
THP-1 lysate
Primary antibody: Anti FLAG, MAb /
2nd antibody: Anti Mouse IgG (H+L),
rabbit, IgG Whole,
POD conjugated
(012-23641)
Lane 1: x 5,000
Lane 2: x 10,000
Lane 3: x 20,000
Lane 4: x 40,000
Sample: mixture of FLAG-BAP and
FM3A cell lysate
Primary Antibody: Anti FLAG,
Polyclonal Ab
2nd antibody: Anti Rabbit IgG (Fc),
mouse, POD
conjugated
(#010-23941)
Lane 1: x 2,500
Lane 2: x 5,000
Lane 3: x 10,000
Lane 4: x 20,000
Grade Description Package
Size
Wako Cat. No.
for
Immunochemistry
Anti Mouse IgG (H+L), Rabbit, IgG Whole, Peroxidase Conjugated 300 μL 012-23641
1 mL 018-23643
Anti Rabbit IgG (Fc), Mouse, Peroxidase Conjugated 300 μL 010-23941
1 mL 016-23943
Anti Rabbit IgG (H+L), Goat, F(ab')2, Peroxidase Conjugated 1 mg 013-17941
Anti Mouse IgG (H+L), Rabbit, IgG Whole, Alkaline Phosphatase Conjugated 1 mg 018-18091
Anti Rabbit IgG (H+L), Goat, IgG Whole, Alkaline Phosphatase Conjugated 1 mg 011-18101
Anti Mouse IgG (H+L), Goat, IgG Whole, Biotin Conjugated, affinity purified 1 mg 010-14031
Anti Rabbit IgG (H+L), Goat, IgG Whole, Biotin Conjugated, affinity purified 1 mg 013-14021
Anti Mouse IgG (H+L), Rabbit, IgG Whole, FITC Conjugated 1.5 mg 012-17531
Anti Rabbit IgG (H+L), Goat, IgG Whole, FITC Conjugated 1.5 mg 011-17621

4. Multi Capture HRP - Antibody Detection Reagent
This is a reagent kit binding to Fc region of antibodies to detect antibodies.
Chemiluminescence can be used for detection because the capture reagent is
HRP-labeled. Mouse IgG1 may be difficult to detect. In that case, use the
Enhancer Reagent (for mouse) provided in the kit.
Features
Almost all antibodies can be detected.
      (Mouse IgG1 and Goat IgG may be difficult to detect.)
Sensitivity is similar or higher than that existing secondary antibodies.
Low lot-to-lot variation

Application 1 Application 2
Primary Ab: Anti FLAG, rabbit, polyclonal Ab
(0.5 μg/mL)
2nd Antibody:
1: Multi Capture HRP ( x 5,000)
2: Multi Capture HRP ( x 10,000)
3: Multi Capture HRP ( x 20,000)
4: HRP conjugated Anti rabbit IgG, Polyclonal
Ab ( x 10,000)
Primary Ab: Anti FLAG, mouse IgG1, MAb (1 μg/mL)
2nd Antibody:
1: Multi Capture HRP ( x 2,000)
2: Multi Capture HRP ( x 2,000) +
Enhancer Reagent for Mouse
3: Anti Mouse IgG (H+L), Rabbit, IgG Whole,
POD Conjugated ( x 5,000)
(Wako Cat. #018-23643)

Kit Contents
10 tests (Wako Cat. #291-71801) 40 tests (Wako Cat. #297-71803)
(1) Capture Reagent 50 μL 200 μL
(2) 20 x Reaction Buffer 10 mL 400 mL
(3) Enhancer Reagent for Mouse 50 μL 200 μL
∗The package size is based on the use when capture reagent is diluted to 1 : 2,000.

Grade Description Package Size Wako Cat. No.
for Blotting Multi Capture HRP 10 tests 291-71801
40 tests 297-71803

5. Immunoenhancer - Accelerator of Antigen-Antibody Reactions
This product is an accelerator to optimize antigen-antibody reaction for Western blotting, dot blotting, enzyme-linked
immunosorbent assay (ELISA). Immuno-enhancer aids in obtaining high signal-to-noise (S/N) ratio.
Features Procedure
Enhances Immunoassay signals, effectively
High S/N Ratio
Applicable to Ready-to-Use Antibody Diluent


Kit contents
for 2 assays∗ for 10 assays. for 40 assays.
Reagent A 10 mL 50 mL 200 mL
Reagent B 10 mL 50 mL 200 mL
∗Each package size shows an assay number when 5 mL each is used
per assay.
Application 1 Application 2 Application 3
A549 cell Lysate 5 g (x1) or 10 g (x2) were
subjected to SDS-PAGE and blotted onto
nitrocellulose membranes. They were blocked
and Western blot.
3% skim milk in TBS-T solution was used as
the control.
Antibodies:
Primary Antibody: Anti EB1, Rabbit (1:500),
2 hour-reaction
Secondary Antibody: HRP conjugated Anti
rabbit IgG (1 : 7000),
1 hour-reaction
Exposure time: 10 seconds
HeLa Cell Lysate, diluted by 1/1, 1/2, 1/4 and
1/8 were subjected to
SDS-PAGE and blotted onto a membrane,
blocked, followed by Western blotting. TBS-T
solution was used as the control.
Antibodies:
Primary Ab: Anti Actin, Goat (0.5 g/mL),
1 hour-reaction
Secondary Ab: Anti goat-IgG-HRP
(1:10,000 dilution),
1 hour-reaction
Exposure time: 5 minutes
Cell extract was separated by 12.5% SDS-PAGE,
transferred onto PVDF membrane, followed by Western
blotting the primary antibody diluted with Immuno-enhancer
Reagent A
. 20 mM Tris-HCl (pH 7.5)-0.15 M NaCl-0.1%
NaN3 containing 1% BSA was used as the control. The
secondary antibody was diluted with 20 mM Tris-HCl (pH
7.5)-0.15 M NaCl-0.05% Tween 20. Generally, LC3-I form is
shown in Atg- cell and LC3-II is mainly detectable in Atg7+.
To detect LC3-I in Atg7+, it is necessary to lengthen the
exposure time. By using Immuno-enhancer, achieve LC3-I
detection, even if the shorter exposure time.
(Data was provided by Dr. Takashi Ueno, Dep. of
Biochemistry, Juntendo Univ. School of Medicine.)
Grade Description Wako Cat. No. Package Size
for Blotting   Immuno-enhancer 2 assays 294-68601
10 assays 290-68603
40 assays 298-68604
  Immuno-enhancer Reagent A 200 mL 091-05811
  Immuno-enhancer Reagent B 200 mL 098-05821

6. Stripping Reagent
Stripping Solution is used to remove primary and secondary antibodies from a Western Blot membrane. Stripping is useful when several proteins are investigated on a sheet of membrane.
Usage
Wash a membrane reacted with a labeled antibody using TBS-T or PBS-T,
and shake the membrane with Stripping Solution for 10 min. at RT.

Application
Sample: Mixture of 1 and 2
1. Transthyretin Variant (L55P), Human, recombinant (Wako Cat. #203-17321)
2. Glutathione S-Transferase
Membrane: PVDF Membrane; Blocking: 0.5% BSA
Labeled Antibody: Anti 6x His, MAb (9C11), HRP conjugated (Wako Cat.#010-23181) 4 mL
Chemiluminescent reagent: ImmunoStar LD (Wako Cat. #296-69901) Exposure time: 1 second

Wash the membrane with TBS-T

Incubate the membrane on an orbital shaker with Stripping Solution for 10 minutes

Wash the membrane with TBS-T

Blocking: 0.5% BSA
Labeled Ab: Anti Glutathione S-transferase, MAb, Peroxidase Conjugated
Chemiluminescent reagent: ImmunoStar LD (#296-69901); Exposure time: 1 second
Grade Description Package Size Wako Cat. No.
for Blotting Stripping Solution 500 mL 193-16375

7. Coloring Reagents
POD Immunostain Set
This is a sensitive enzyme-immunostaining kit of horseradish peroxidaselabeled
immunoglobulins on nitrocellulose membrane. Phenol is oxidized
by reaction of phenol and hydrogen peroxide in the coloring reagent with
POD, and blue-purple diformazan is generated in proportion to POD
activity, in the presence of NADH and NTB. Antigen is detected by this
reaction.
Kit Contents
NTB solution 250 mL x 1 bottle
NADH for 20 mL x 12 bottles
Substrate solution 13 mL x 1 bottle
Grade Description Package Size Wako Cat. No.
for POD Staining POD Immunostain Set 20 mL x 12 299-18841

ABC Solution
This is Streptavidin-Biotin-Peroxidase Complex Solution (ABC Solution). The solution reacts with biotin-labeled secondary
antibodies to form antigen-primary antibody-secondary antibody-ABC complex. Then substrate is added to detect protein
bands. Antigen can be detected with high sensitivity because secondary antibody binds to multiple peroxidase via
streptavidine.
Grade Description Package Size Wako Cat. No.
for Immunohistologic ABC Solution Staining 10 mL 017-15881

BCIP/NBT Solution
This is mixture solution of 5-Bromo-4-chloro-3-indolyl-phosphate (BCIP) and Nitroblue Tetrazolium (NBT). BCIP and NBT
generate an insoluble, dark blue-purple colored product by alkaline phosphatase. This reaction can be used for detection by
Western blotting. The solution is prepared as a substrate for coloring in advance at a concentration suitable for
immunoblotting.
This product is inapplicable to a microwell-EIA and immunohistostaining.

Grade Description Package Size Wako Cat. No.
for Biochemistry BCIP / NBT Solution 100 mL 022-16231

8. ImmunoStar LD Chemiluminescence Reagent
High-sensitive chemiluminescent detection in a short time

ImmunoStar LD (Long Detection), which is designed for a simple and highly
sensitive immunoblotting, utilizing detection by enhanced chemiluminescence
using our unique luminol derivative L-012 as the substrate can be detect
labeled HRP.
Signal of Western blotting can be detected with high sensitivity due to
combination with unique enhancer. Also, it has long duration of luminescence.
Principle
L-012 is a luminol derivative, and dianion generated from H2O2 shows chemiluminescence.

Features
High Sensitivity (10-14; femto gram level)
Long term duration of chemiluminescence (CHL) (24 hours)
Low background (a high ratio of S/N)
Easy preparation (Prepare Luminescence Working Solution by mixing Soln. A and B at a ratio of 1:1)

Grade Description Package Size Wako Cat. No.
for Blotting ImmunoStar LD 200 cm2 296-69901
1,000 cm2 292-69903
2,000 cm2 290-69904
: Not available for sales in the US and Europe.
References
1. Obana, N., et al. : Mol. Microbiol., 77, 1416 (2010)
2. Fukumoto, H., et al. : J. Neurosci., 30, 11157 (2010)
3. Hyakkoku, K., et al. : Neuroscience., 171, 258 (2010)
4. Obana, N., et al. : J. Bacteriol., 193, 4417 (2011)
5. Kurotani, R., et al. : J. Biol. Chem., 286, 19682 (2011)
ImmunoStar LD
High-sensitive detection in a short time
A good result with low background can be obtained by exposure for several seconds to minutes.



A mixture of synthesized Aβ, 37, 38, 39, 40 and 42 was subjected to
SDS-PAGE using Tris-Tricine Urea gel that can separate Aβ molecule
species, then transferred to PVDF membrane and boiled up. Western
blotting was performed using an antibody specifi c to N terminal of
human Aβ.
(Data was provided by Dr. Tomita and Osawa, Department of Neuropathology and Neuroscience,
Graduate School of Pharmaceutical Sciences, The University of Tokyo)

Duration of Luminescence
Examination of dillution factor of secondary antibody.
Related Product
We recommend the following reagent when the sensitivity of ImmunoStar LD is higher than that you expect.
Grade Description Package Size Wako Cat. No.
for Blotting ImmunoStar Reagents for 1,000cm2 295-55201
for 5,000cm2
291-55203

D. MS Analysis

1. Reagents for Sample Pretreatment
     a. Proteome Preparation Reagent

This product easily removes mucopolysaccharides and nucleic
acids from extracted samples of plants, animals and
microorganisms. As a result, the viscosity is decreased and the
fi gure of electrophoresis is improved.
Description Package
Size
Wako Cat. No.
Proteome Preparation Reagent 100 μL 164-22691

     b. Hydrophobic Protein Preparation Reagent
Hydrophobic Protein Preparation Reagent is used to accelerate solubilization of hydrophobic proteins and to obtain twodimensional
electrophoretic spots of proteins that cannot be isolated by a conventional method. It is also used to reduce
the effects of lipids and nucleic acids contaminated in soluble proteins and obtain clearer spots.
Description Package
Size
Wako Cat. No.
Hydrophobic Protein Preparation Reagent 0.2 mL 087-08891
0.5 mL 083-08893

2. Matrix for MALDI-TOFMS Analysis
     MS Analysis
High-purity Matrix for MALDI-TOFMS analysis
CHCA, SA and DHB

Features
1. High purity matrix is mixed with a test sample and used for proteome analysis by mass spectrometry (MALDI-TOFMS).
2. High purity matrix gives good mass spec reading because of its high purity by recrystallization).

Effects of recrystallization of α-Cyano-4-hydroxycinnamic Acid (CHCA)
When comparing with MALDI-TOFMS data for commercial CHCA, it was found that recrystallized CHCA (Wako Cat. No. 037-19261) gives
clearer mass spec readings with less background noise. (These data were provided by Dr. Wada Y. at Osaka Medical Center, Japan)

Product List
Grade Description Package
Size
Wako Cat. No. Storage
for proteome research α-Cyano-4-hydroxycinnamic Acid [CHCA] 50 mg x 5 037-19261 2-10℃
Sinapic Acid [SA] 50 mg x 5 192-13361
2,5-Dihydroxybenzoic Acid [DHB] 50 mg x 5 044-29101

3. In-gel Digestion Enzymes
     Lysyl Endopeptidase, MS Grade
Among the most important techniques in proteome analyses is the in-gel digestion of protein
spots/bands that have been resolved by electrophoresis using digestive enzymes, such as
trypsin and lysyl endopoptidase. Proteins can be identifi ed by mass spectrometry analysis of
the peptides produced by in-gel digestion, and further information regarding posttranslational
modifi cations can be obtained.
Lysyl Endopeptidase, Mass Spectrometry Grade is a lyophilized product that retains
sufficient activity for in-gel digestion and packed in very small quantities for convenience
purposes.

Features
1. High specifi city and efficiency of protein digestion allow for easy database searches by peptide mass.
2. Improved cleavage at lysine residue and increase in the number of peptides are obtained by combination with trypsin.
3. Packed in very small quantities in accordance with common usage guidelines, so sufficient activity is retained for in-gel digestion.
Comparison of In-gel Digestion Using Trypsin (Tp), Lysyl Endopeptidase (Lep) and Lep Combined with Tp (Lep +Tp)
BSA band (100 ng) resolved by SDS-PAGE was in-gel digested with Tp, Lep and Lep +Tp and analyzed by MALDI-TOFMS.
The fi gure shows the individual mass spectra. The evaluation of these peptidases is summarized in the table.
Table: Comparison of Tp, Lep and Lep +Tp
These results indicate there are very few missed cleavages obtained by Lep digestion. When Tp is used concomitantly with Lep, missed
cleavages decrease and the number of identifi ed peptides increase compared to when only Tp is used.
Tp Lep Lep +Tp
Cleavage site C terminal of Arg and Lys C terminal of Lys C terminal of Arg and Lys
Missed cleavage (Rates of missed cleavage)∗1 Many (8 %) Very few (0 %) Few (3 %)
No. of identifi ed peptides 17 19 22
∗1 The value resulted from subtracting the coverage obtained when database searches were performed with Missed cleavage 0 from that obtained when
performed with Missed cleavage 1. "Coverage" is the percentage of peptides obtained after in-gel digestion in the whole sequence.
Figure: Comparison of the mass spectra of (a) Trypsin (Tp) and (b) Lysyl Endopeptidase (Lep) +Tp
The peaks at m/z 2000 were obtained after digestion with Lep + Tp, but not Tp alone. These results indicate improved sequence c overage.
(Data provided by Dr. Y. Wada, Osaka Medical Center and Research Institute for Maternal and Child Health)
Description Wako Cat. No. Package Size Grade
Lysyl Endopeptidase®, Mass Spectrometry Grade 125-05061 20 μg x 5 for Proteome Research

Related Products
Description Wako Cat. No. Package Size Grade
Trypsin, from Porcine Pancreas, Mass Spectrometry Grade*2 202-15951 20 μg x 5 for Proteome Research
Endoproteinase Asp-N, Sequencing grade 056-05921 2 μg for Sequencing Grade
Endoproteinase Glu-C, Sequencing grade 050-05941 50 μg
V8 Protease [Endoproteinase Glu-C] 164-13982 2 mg for Biochemistry
∗2 : Not available for sale in the US
References
1. Wada, Y., and Kadoya, M.: J. Mass Spectrom., 38, 117 (2003).
2. Shevchenko, A., Wilm, MM., Vorm, O., and Mann., M.: Anal. Chem., 68, 850 (1996).

4. LC/MS Solvents
Liquid chromatography - mass spectrometry (LC/MS) is widely used in various fi elds including
biological, food, and environmental analyses. In particular, recent breakthroughs in the
development and upgrades of device interfaces have led to the use of LC/MS in microanalyses of
environmental pollutants and chemical metabolites, etc.
Following products are ideal LC/MS reagent to analyze trace components.

Description Wako Cat. No. Package Size Features Specification
Acetic Acid 014-20063 1 mL x 5 A . Suitability test for LC/MS analysis performed
. Reduced background noise
Assay (HPLC): 99.5+%
Absorbance (1 → 4,250 nm): max. 0.50
Absorbance (1 → 4,254 nm): max. 0.10
Fluorescence test: to pass test
Suitability for LC/MS analysis: to pass test
018-20061 50 mL
Acetonitrile 016-19854 100 mL . Suitability test for LC/MS analysis performed
. Guarantees noise level at m/z 50~2,000
. Use of aluminum caps
Reduced risks of slight amounts of
contaminants from plastic caps
Assay (cGC): 99.8+%
Density (20°C): 0.780 ~ 0.783 g/mL
Fluorescence test: to pass test
Suitability for LC/MS analysis: to pass test
012-19851 1 L
018-19853 3 L
Formic Acid
(abt. 99%)
063-04533 1 mL × 5 A . Suitability test for LC/MS analysis performed
. Reduced background noise
Assay (HPLC): 99.5+%
Solubility in water: to pass test
Absorbance (1 → 4,254 nm): max.1.00
Fluorescence test: to pass test
Suitability for LC/MS analysis: to pass test
067-04531 50 mL
0.1 vol% Formic
Acid-Acetonitrile
062-04721 1 L . Suitability test for LC/MS analysis performed
. Ready-to-Use eluent
Absorbance (200-400 nm): to pass test
Fluorescence test: to pass test
068-04723 3 L Water: max. 0.05%
068-04723 3 L
Methanol 132-14524 100 mL . Suitability test for LC/MS analysis performed
. Guarantees noise level at m/z 50-2,000
. Use of aluminum caps
Reduced risks of slight amounts of
contaminants from plastic caps
Assay (cGC): 99.7+%
Density (20℃): 0.789-0.792 g/mL
Fluorescence test: to pass test
Suitability for LC/MS analysis: to pass test
138-14521 1 L
134-14523 3 L
Ultrapure Water 214-01301 1 L . Decreased total organic carbon levels
. Guarantees the absorbance and fl uorescence
tests
. Use of specially processed glass containers /
aluminum caps
Density (20℃): 0.997 ~ 0.999 g/mL
Refractive index nD20: 1.332 - 1.334
Absorbance (210-400 nm): max. 0.01
Fluorescence test: to pass test
210-01303 3 L Total organic carbon (TOC): max. 4 ppb
210-01303 3 L

Related Products
Description Wako Cat. No. Package Size Features Specification
Wakopak®
MS-5C18GT
001-00030 2.0 mmφ x 50 mm . The glass lined inside wall of the stainless column allows
maximum inactivation
. Excellent peak shapes and recovery rates in analyses of trace
components within biological samples
DuPont
2.0 mmφ x 100 mm
2.0 mmφ x 50 mm

5. MALDI-MS Calibrants
For MALDI-MS Calibration
Peptide standards for matrix-assisted laser desorption ionization (MALDI) mass spectrometry used extensively for analysis
of biological polymers such as proteins are highly purifi ed meeting the requirements of MALDI-MS. They are available in
step sizes of 500 in molecular weight for selection of calibrants corresponding to your sample.
Features
1. High Purity
2. Appropriate calibrant for your sample
3. Cost-conscious 2 nmol package
[MS Chart Example 1] Example of Use
[MS Chart Example 2] [MS Chart Example 3]
Description (M+H)+ Wako Cat. No. Package Size
Angiotensin II (Human)
<MALDI-MS Calibrant>
①1046.5423 019-22931 2 nmol x 1 bottle
015-22933 2 nmol x 5 bottles
[D-Arg1, D-Pro2, D-Trp7,9, Leu11]-Substance P
<MALDI-MS Calibrant>
②1497.8483 017-22971 2 nmol x 1 bottle
013-22973 2 nmol x 5 bottles
Apamin
<MALDI-MS Calibrant>
③2026.8944 016-22941 2 nmol x 1 bottle
012-22943 2 nmol x 5 bottles
PAMP (Rat)
<MALDI-MS Calibrant>
④2477.3642 160-24251 2 nmol x 1 bottle
166-24253 2 nmol x 5 bottles
ACTH (Human, 1-24)
<MALDI-MS Calibrant>
⑤2932.5885 014-22981 2 nmol x 1 bottle
010-22983 2 nmol x 5 bottles
Adrenomedullin (Human, 22-52)
<MALDI-MS Calibrant>
⑥3574.877 013-22951 2 nmol x 1 bottle
019-22953 2 nmol x 5 bottles
Insulin (Human)
<MALDI-MS Calibrant>
⑦5804.6455 090-05881 2 nmol x 1 bottle
096-05883 2 nmol x 5 bottles


Listed products are intended for laboratory research use only, and not to be used for drug, food or human use.
Please visit our online catalog to search for other products from Wako ; www.e-reagent.com
This brochure may contain products that cannot be exported to your country due to regulations.
Bulk quote requests for some products are welcomed. Please contact us.