KENDRICK LABS CATALOG    

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Call 800-462-3417 or email 2d@kendricklabs.com for a price quote or to receive printed information by mail.

2D ELECTROPHORESIS

MK-1 Mailing Kit This kit contains ten 1 ml tubes of Urea Sample Buffer, ten 1 ml tubes of SDS Boiling Buffer; ten 1 ml tubes of SDS Boiling Buffer minus BME; ten 1 ml tubes of Osmotic Lysis Buffer; ten 100 μl tubes of 10X Nuclease Stock; three 100 μl tubes of Protease Inhibitor Stock Solution and two 100 μl tubes each of Calbiochem phosphatase inhibitor cocktail sets I and II (cat No 524624 & 524625). The compositions of these solutions is given in the Kendrick Labs booklet "Suggestions for Sample Preparation". In addition a Styrofoam mailer, inner carton for radioactive samples and mailing instructions are included.
Price: $250 includes shipping on dry ice.

2D-ES-1: 2-D ELECTROPHORESIS BASIC SERVICE Our standard gel size is perfect for immunoprecipitations, subcellular fractionations, most bacterial preparations and many proteomics applications.  One Coomassie blue stained, dried 2-D gel of dried dimensions 15 cm wide x 13 cm long, is returned for each sample along with a pH gradient plot and a method description sheet suitable for reports or publications. The example below is from an Escherichia coli sample prepared in our Urea sample buffer (left) and SDS Boiling Buffer (right) run identically on a pH 4-8 IEF gradient and 10% acrylamide slab gel. The great advantage of SDS buffer is ease of sample preparation. Molecular weight standards (220, 94, 60, 43, 29, and 14 kDa) appear as bands on the right side of the gel labeled with press-on numbers. One isoelectric point marker, added to each sample as an internal standard, is marked with an arrow. Our standard IEF tube gel contains 2% pH 3.5-10 mix4L or 2% 4-8 mixed ampholines; transparency drying (STP-8) is standard. The duplicate gel rate is used for duplicate gels with different stains and also for blots, but not for different gel conditions.  More photos of example gels. See 2D-ES-5 for our large format option.
Price:

2D-ES-1: 1 sample, $175

2D-ES-1A: 2-3 samples, $165 each

2D-ES-1B: 4-8 samples, $155 each

2D-ES-1C: 9-20 samples, $146 each

2D-ES-1D: 21-200 samples, $140 each

2D-duplicate: $85 each

More than 200 samples - Inquire

CUSTOM PH GRADIENTS are available using ampholines of pH 3.5-10, 2.5-5, 4-6 and 7-9. For basic proteins with pI's greater than 9, non-equilibrium pH gradient electrophoresis is used (NEPHGE, O'Farrell, P. et. al. Cell 12: 1133, 1977). Buffers and electrodes are reversed and the pH gradient is created using pH 8-10.5 ampholines. Indicate which custom pH gradient you need on the back of the sample ID form. (No extra charge)

2D-ES-2: 2D ELECTROPHORESIS WITH PEPTIDE SLAB GELS. The 16.5% peptide slab gels of Schagger and von Jagow (Anal. Biochem. 166: 368, 1987) are useful to resolve low MW proteins (3 - 14 kDa). In general it is better to load low MW proteins heavily (10 μg/spot).  Note that ampholines may interfere in the MW range 2-6 kDa. 
Price: Add $20 per gel.

2D-ES-3: NATIVE 2D ELECTROPHORESIS For native 2D the urea and IGEPAL (NP-40) are omitted from the IEF tube gel; SDS and BME are omitted from the 2nd dimension slab gel. No denaturing agents are used in the sample buffer. Many proteins are insoluble under these conditions and don’t resolve. However, occasionally it’s very useful for soluble proteins in serum and CSF.
Price: Same as 2D-ES-1. See also 2D-ES-6: Blue Native Gel Electrophoresis.

2D-ES-4: 2-D ELECTROPHORESIS WITHOUT SULFHYDRAL REAGENT  Same as 2D-ES-1 except 2-mercaptoethanol and dithiothreitol are omitted from sample buffer, internal standards, and all 2-D solutions.
Price: Same as 2D-ES-1.

2D-ES-5  LARGE FORMAT 2D ELECTROPHORESIS  The larger size gives better resolution for proteomics applications; heavier loads may be applied to obtain more material for mass spectrometry. One Coomassie blue-stained, dried 2D gel of dimensions 20 cm wide by 20 cm long is returned for each sample along with a pH gradient plot and a method description sheet.  Our standard IEF tube gel contains 2% pH 3.5-10 mix4L or 4-8 mix ampholines. Molecular weight standards (220, 94, 60, 43, 29, and 14 kDa) appear as bands on the right side of the gel labeled with press-on numbers. One IEF marker, added to each sample as an internal standard, is marked with an arrow. Transparency drying (STP-8) is standard. See above for Custom pH gradients. Typical protein loads are 100 (silver staining) or 600 μg (Coomassie blue) in 25-150 μl as shown in images below.
Price:

2D-ES-5: 1 sample $245

2D-ES-5A: 2-3 samples $235 each

2D-ES-5B:  4-12 samples $225 each

2D-ES-5C: 13-200 samples $215 each 

LF-Dup (Large Format 2D duplicate gels): $100 each 

More than 200 samples - Inquire.  

Large Format 2D Gel Examples, silver versus Coomassie staining. A side by side comparison of 2D gels loaded with same sample and stained with Coomassie blue (left, 600 µg sample load) and with silver (right, 100 µg sample load). The silver stain is at least 10x more sensitive than Coomassie blue stain even though less is loaded.
 

2D-ES-6: Blue Native Polyacrylamide Gel Electrophoresis (BN-PAGE) is a charge shift method developed by Schagger and von Jagow (Anal. Biochem. 199: 223-231, 1991) for isolation and characterization of large multi-protein complexes (MPCs) in their native state. Binding of Coomassie blue G-250 in the absence of denaturing detergents allows for first dimension separation according to the size and shape of the complex on 4-13% gradient gels.

Coupling first dimension BN-PAGE with a second dimension of SDS-PAGE allows for the dissociation of individual subunits based on molecular weight. Further analysis by Western blotting or mass spectrometry can be performed to identify specific subunits within MPCs. We currently offer both BN-PAGE as the first dimension only, or BN-PAGE followed by SDS-PAGE as a second dimension. BN/SDS PAGE has proven a valuable tool for studying the functional proteomics of mitochondrial protein complexes (Methods 26: 327-334, 2002), membrane protein complexes (Anal. Biochem: 217: 220-230, 1994), and whole cell lysates (Molecular and Cellular Proteomics: 3.2: 176-182, 2004). Please call to discuss before sending samples for BN-PAGE. 
Price:

2D-ES-6: 1 sample $175

2D-ES-6A: 2-3 samples $165 each

2D-ES-6B: 4-8 samples $150 each

2D-ES-6C: 9-20 samples $140 each

2D-ES-6D: 21-200 samples $130 each

BN-PAGE (1 D gel only): $110 first sample

BN-PAGE-A: $50 each additional

2D-ES-7: CARBAMYLATED CARBONIC ANHYDRASE pI MARKERS This protein was purchased from Sigma-Aldrich, progressively carbamylated on lysine residues by heating in urea buffer for various times, and calibrated for pI. Each spot in the beads-on-a-string pattern differs from the next by a single charge; measured pI values are shown in the table. A pH gradient standard curve may be determined from the train of charge isomers added an an internal standard for calculation of pIs for unknown proteins. Note that these pIs are not absolute values but are for conditions of 9 M urea and 22^o C. The arrow marks an internal standard protein, tropomyosin, MW 33, 000, pI 5.3. The right-most spot is 1, pI 7.3.

Price: Add $16 per gel.

  2DE/Top   Mass Spectrometry   Staining   Blotting   Radioactivity   Sample Preparation   Misc

2D-ES-9: MANUAL COMPARISONS of 2D PATTERNS FOR DIFFERENCES. We find that HBC (Human Brain Comparison) is a reliable method for finding changes in 2D patterns. In this method two experienced analysts independently compare stained gel pairs on a light box for differences. Results are presented as color coded outlines of spots on  transparent overlay covering the  gels. Free duplicate gels are run to confirm differences for samples scheduled for manual comparisons.
Price:

2D-ES-9: $165 per Standard Format pair compared

2D-ES-9LF: $195 per Large Format pair compared

2D-ES-10: COMPUTERIZED COMPARISONS OF 2D GEL PATTERNS FOR DIFFERENCES.   This analysis includes scanning with a calibrated densitometer and computerized analysis with SameSpots software v4.0 (2010) from Non-Linear Dynamics. Requires prior 2D electrophoresis but free duplicate gels are run and analyzed to confirm differences for samples scheduled for computerized comparisons. Generally 400-800 spots are quantified per Standard Format gel; 800-1200 spots are quantified per Large Format gel. Spot density values are expressed as spot percentages (individual spot density as a percentage of total density in all spots analyzed) to normalize for differences in sample loading or staining.

Results are presented in a complete report including a summary table showing spot number, pI, MW, ratios (fold difference) and p values as a second measure of difference. The final report also includes figures of images showing numbering, montage images for every differing protein spot, methods and pH gradient plot. A CD containing data and image files is included in the package. Note that our reports focus on the protein differences between samples. Picking protein spots later for identification by mass spectrometry is straightforward. Just send us a list of spot numbers are we'll take it from there.
 

Price: Inquire for project-specific quote.

2D-ES-10 (Standard Format): duplicate gels analyzed and averaged per sample. Per comparison, $450.

2D-ES-10LF (Large format): duplicate gels analyzed and averaged per sample. Per comparison, $500.

2D-ES-12 QUANTIFICATION OF PROTEINS RESOLVED BY 2-D ELECTROPHORESIS. For clients knowing which proteins need to be quantified. Scanning (2D-ES-13) is included. Data is reported in integrated spot density in density units, or as a percentage of total image density. Spots are not matched from gel to gel.
Price:

2D-ES-12: 1-12 spots $85

2D-ES-12A:13-50 spots $145

2D-ES-12B:1-200 spots $180

2D-ES-12C:1-400+ spots $285 

2D-ES-13: LASER DENSITOMETRY. Stained gels and x-ray films are digitized with our laser densitometer from Molecular Dynamics. The densitometer is linear over the range 0-3.0 optical density units as verified by calibrated filters. Narrow O.D. ranges can be expanded to full scale (8 bit, 0-255) if required. Scanning resolution is 100 microns/pixel; the final black & white tif images are 3.3 Mbytes each for standard format and 7 Mbytes for large format.

Price: $25/scan.

2D-ES-13A: FLUORESCENT DENSITOMETRY with a Bio-Rad Molecular Imager ProPlus. Images are scanned at 100 micron resolution.

Price: $35/scan

2D-ES-13B: Quantitative densitometry with a GE Healthcare Image Scanner III calibrated to be linear from 0-2.3 OD using Melles Griot NIST-calibrated filters. Images are scanned at 84 micron resolution.

Price: $25/scan

2D-ES-14: ELECTRONIC PHOTOS AND FILES. The 10 mB black & white files obtained from laser densitometry are too big to email and sometimes don't look right by eye. Our Afga Arcus II and HP desktop scanners give excellent color images in tif format that are useful for publication and emailing (jpg format) The price includes tif and jpg image files on a CD mailed with the dried gels, and jpg images emailed to provide previews of 2D patterns. 
Price: $15 each

2DE/Top   Comparisons   Staining   Western blotting   Radioactivity   Sample Preparation  Misc

Protein Identification by Mass Spectrometry

We have considerable experience in sending 2D gel proteins to university and commercial core facilities for identification by mass spectrometry fingerprinting (also called peptide mass matching) and for amino acid sequencing.  Turnaround time for mass spectrometry is usually about two weeks.  We highly recommend Columbia University Protein Core Facility  Contact Dr. Mary Ann Gawinowicz, the facility director, for more information. tel:  212-305-3631, email: mag4@columbia.edu

2D-ES-19 Spot cutout for mass spectrometry Fingerprinting or sequencing  We do this manually and are obsessively careful. The cutout spots are placed in Eppendorf tubes for express mail to the core facility of your choice; our default is Columbia University. The cost includes a letter to the core listing the spots, plus a copy of the letter for you along with an image or original gel showing spot locations. We make sure that each spot has a unique identifier.

Price: $20 per spot

2D-ES-15  MASS SPECTROMETRY FINGERPRINTING (peptide mass matching): This technique is used to identify proteins from Coomassie blue or special silver-stained gels. The protein must be known and in the databases. The cutout polypeptide spot is digested with an enzyme of known specificity, usually trypsin, and the resulting peptide mixture, without separation, is analyzed by MALDI (matrix-assisted laser desorption ionization) MS. The peptide masses obtained by MS are entered into a search program which scans the database, NCBI or Genpept, to find a match. This method will not produce sequence data from the protein, only a unique set of peptide masses enabling identification. The mass spectrum will usually give 25-60% sequence coverage (number of amino acids in identified peptides/number of amino acids in whole protein).
Price for 2D-ES-15:  $190 per polypeptide spot including database searching by hand, and invoicing through Kendrick Labs.

Search programs used at the Columbia Protein Core Facility are ProFound at http://prowl.rockefeller.edu/prowl-cgi/profound.exe  MS-Fit at http://prospector.ucsf.edu , and Mascot at www.matrixscience.com 

References for peptide mass mapping
1.  Shevchenko, A.,  Jensen, O.N., Podtelejnikov, A.V., Sagliocco, F., Wilm, M., Vorm, O., Mortensen, P., Boucherie, H., and Mann, M., Proc. Natl. Acad. Sci. U.S.A. 93: 14440-14445 (1996)
2. Jensen ON, Podtelejnikov, AV, and Mann, M. Identification of the components of simple protein mixtures by high-accuracy peptide mass mapping and database searching. Anal Chem. 69:4741-50 (1997).
3. Henzel WJ, Watanabe C, Stults JT. Protein identification: the origins of peptide mass fingerprinting.  J Am Soc Mass Spectrom. 9:931-42 (2003).

2D-ES-18  MASS SPECTROMETRY using LC/MS/MS: This powerful method is used to identify proteins or to determine the unknown amino acid sequence of peptide fragments of the protein of interest cut from Coomassie blue or special silver stained gels.  This method is recommended for spots or bands which might contain mixtures, for spots which could not be identified by MALDI, and for samples from species with incomplete genomes.

Protein spots are cut from Coomassie blue or special silver-stained gels and digested with trypsin. The digestion mixture is separated on an LC Packings nano-lc on which the detector outlet is connected directly to the nanospray source of a Micromass Q-Tof mass spectrometer. Peptides are eluted at a flow rate of 200 nl/min and are scanned as they enter the source. When a peptide is detected, the Q-Tof is programmed to switch to MS/MS mode, which means that the eluting peptide is fragmented by means of an applied collision energy and the resulting ions scanned for several seconds. When the programmed time for MS/MS is done, the Q-Tof switches back to MS mode and resumes scanning the eluting peptides. Because peptides fragment at the peptide bond in a predictable way, the fragmentation pattern can be used to deduce sequence information. The ions resulting from fragmentation along with the mass of the intact peptide can be used to search a database and identify the protein. Proteins which show a good MALDI pattern but cannot be identified by Mass Spectrometry Fingerprinting (2D-ES-15) are candidates for identification by LC/MS/MS. Generally, LC/MS/MS is not more sensitive than MALDI, but the data is acquired differently and can be more informative.
Price for 2D-ES-18 LC/MS/MS:  $370 per polypeptide spot.

Reference for protein identification by LC-MS/MS
1. Hernandez, P, Muller, M, and Appel, RD. Automated protein identification by tandem mass spectrometry: issues and strategies. Mass Spectrom Rev. 25: 235-254 (2005).
 

2D-ES-20  PHOSPHORYLATION SITE IDENTIFICATION BY MASS SPECTROMETRY.  We outsource this state-of-the-art service to the experts at the Laboratory for Proteomic Mass Spectrometry at the UMass Med School. See:  www.umassmed.edu/proteomic/Leszyk/index.aspx for more information. Contact Dr. John Leszyk, Tel: 508-856-7533  Email: John.Leszyk@umassmed.edu   Note that the success of phosphoprotein analysis depends on the type of protein and the extent of phosphorylation. In some cases additional work may be required. Note that the success of phosphoprotein analysis depends on the type of protein and the extent of phosphorylation. See our Feb 09 Newsletter for a brief literature review and advice on amount of phosphorylated protein required.
Price 2D-ES-20A: $520/polypeptide spot

2D-ES-17  N-TERMINAL AMINO ACID SEQUENCING BY EDMAN DEGRADATION. Typically done after 2D electrophoresis and PVDF transblotting to determine if a protein is clipped. The protein must be loaded heavily enough to see with Coomassie blue on the blot; the N-terminus cannot be blocked.  Note that bacterial proteins are seldom blocked at the N-terminal while mammalian proteins are often blocked, up to 60-70% of the time. This is older technology but useful in some cases, for example to see if a protein has been clipped by a protease. 
Price:  $170 setup plus $25 per cycle

2D Gels/Top   Comparisons   Mass Spectrometry   Blotting   Radioactivity   Sample Preparation   Misc
 

STAINING/DRYING PROCEDURES

STP-1: COOMASSIE BLUE STAINING. We use Coomassie brilliant blue R250 as our standard staining technique (O'Farrell, P. J. Biol. Chem. 250: 4007, 1975). This method, performed according to SOP L-1510, involves an alcohol/acetic acid fix, an acetic acid rehydration and an acetic acid destain. Generally, 1 µg of purified protein gives a nice spot on a 2D gel (see our internal standard) and 0.1 ug is visible. Although not as sensitive as silver staining, this method gives quantifiable results. (Burgess-Cassler et. al. Clin. Chem. 35: 2297, 1989.) The linear range for plots of stain density versus ng protein is broad and the method is reproducible. This stain is compatible with mass spectroscopy. 

Price: No charge (This is our standard stain).

STP-2: SILVER STAINING.  Our silver stain is based on the ammoniacal silver/formaldehyde method of Oakley et. al. (Anal. Biochem.105: 361, 1980.) This method involves preliminary glutaraldehyde treatment of the slab gel to fix proteins by cross linking. The pre-treatment also adds glutaraldehyde side chains to the proteins, increasing sensitivity since these groups are sites for silver deposition (Dion A, and Pomenti A, Anal. Biochem. 129: 390, 1983). We find this method to be about 10 times more sensitive than Coomassie blue staining, depending on the protein, even though less protein is loaded. Generally, 50 ng of purified protein gives a highly visible spot on a 2D gel (see our internal standard). However, some proteins that are detectable with Coomassie don't stain at all with silver. Although very sensitive, this method gives semi-quantitative rather than quantitative results. The linear range for plots of stain density versus ng protein varies widely from protein to protein and also because day to day variability in stain and background intensity is observed. Silver staining is not compatible with mass spectroscopy fingerprinting.
Price:

STP-2 (Standard Format): $32 per gel

STP-2LF (Large Format): $42 per gel

STP-3: SPECIAL SILVER STAINING FOR MASS SPECTROSCOPY ANALYSIS.  This special silver stain (O’Connell and Stults, Electrophoresis 18: 349-359, 1997) omits the glutaraldehyde step and is compatible with mass spectroscopy fingerprinting. It often negatively stains proteins and so is not appropriate for computerized analysis.  However, the pattern can be matched to our regular stain without difficulty. Note however, that silver deposition interferes with the enzymatic digestion required for mass spectrometry for most mammalian proteins.  If possible use the less sensitive but more reliable Coomassie stain. 
Price:

STP-3 (Standard Format): $32/gel

STP-3LF (Large Format): $42/gel

STP-4: SYPRO RUBY FLUORESCENT STAIN. This Molecular Probes stain fluoresces linearly with protein over a range of 100-1000 ng and is compatible with mass spectrometry. See 2D-ES-13A Fluorescent densitometry for imaging.
Price:

STP-4 (Standard Format): $65 per gel

STP-4LF (Large Format): $90 each  

STP-5: Cy dye staining for DIGE analysis
Price: Inquire for quote.

STP-6: EMERALD GLYCOPROTEIN  FLUORESCENT STAIN This Molecular Probes stain is specific for glycoproteins and also compatible with mass spectrometry but isn't very sensitive. See 2D-ES-13A Fluorescent densitometry for imaging.
Price:

STP-6 (Standard Format): $105 per gel

STP-6LF (Large Format): $210 per gel  

STP-7: PRO-Q DIAMOND STAIN FOR PHOSPHOPROTEINS This Invitrogen stain uses a novel fluorophore that recognizes phosphate groups on proteins directly on gels - without antibodies or radioactivity. Although many groups have had success with this stain we cannot guarantee sensitivity or specificity. See 2D-ES-13A Fluorescent densitometry for imaging.

Price: 

STP-7SF (Standard Format): $60 per gel

STP7-LF (Large Format): $75 per gel

STP-8: TRANSPARENCY DRYING. All stained gels are air dried between cellophane sheets unless otherwise requested. Although this treatment adds a day to turnaround, the advantages of easier storage, little or no curling, good color retention, overlay capability, scanning capability, and overhead projection capability offset the time delay. Since cellophane quenches ³⁵S and ³H, gels with these isotopes scheduled for autoradiography or fluorography are paper dried (STP-9).

Price:  No extra charge.

STP-9: PAPER DRYING  This is an optional drying method for stained gels. All Enhance-treated slab gels scheduled for fluorography as well as unstained gels scheduled for autoradiography are dried onto thick filter paper unless otherwise requested.

Price: No  extra charge.

STP-10: WET GELS  In some cases the return of wet gels is requested, for example, for a core facility's automatic spot picker. This is no problem. The wet gels are placed between sheets of thick wet filter paper and the sandwich placed in a ziploc bag. The bag is supported by pressboard sheets and cushioned by bubble wrap prior to express mailing. Gel identification information is written on the ziploc bag. 

Price: No extra charge.

2D Gels/Top   Comparisons   Mass Spectrometry   Staining   Blotting   Sample Preparation   Misc

RADIOACTIVITY PROCEDURES

Kendrick labs has a Radionuclide License (NRC) that can be faxed or emailed to your RSO to ease the shipment of radiolabeled samples. See page 3 for radioisotope mailing instructions.

RP-1: AUTORADIOGRAPHY. Includes 2 film exposures at room temperature using 8" x 10" Kodak BioMax MR film. Exposure times are usually 2 and 4 days.
Price: Add $55 per gel.

RP-2: ENHANCE TREATMENT. For fluorography of low energy emitters such as ³H, ¹⁴C, and ³⁵S. (New England Nuclear reagent and protocol). The gel is soaked in a solution (Enhance) containing a fluorescent intermediate which, at low temperatures, converts beta-particle energy into photons. This treatment reduces exposure times for ¹⁴C and ³⁵S to about 1/8 that of untreated gels. Note that disposal costs for used Enhance add to the cost.
Price:

RP-2 (Standard Format): add $45 per gel

RP-2LF (Large Format): add $65 per gel

RP-3: FLUOROGRAPHY. Requires prior Enhance treatment (RP-2). Includes 2 film exposures at -70ºC using gel 8" x 10" Kodak BioMax MR film. Exposure times are usually 2 and 4 days.
Price: Add $55 per gel.

RP-4: 14C-MOLECULAR WEIGHT MARKERS. These markers from Amersham are added during slab gel electrophoresis so that bands appear on X-ray film after exposure of the gels. These markers have molecular weights of 200, 97.4, 69, 46, 30 and 14.3 kDa.
Price: Add $15 per gel.

RP-6  RADIONUCLIDE DISPOSAL FEE to help cover the high cost of our Radioactive Materials License and disposal fees.
Price: $15 per radiolabeled sample.  

2D Gels/Top   Comparisons   Mass Spectrometry   Staining   Radioactivity   Sample Preparation   Misc

BLOTTING & WESTERN BLOTTING   P-Tyr WB package      P-Ser/P-Thr WB package

BP-1: TRANSBLOTTING ONTO PVDF. PVDF (Immobilon from Millipore Corp.) binds proteins more tightly than nitrocellulose and gives better recoveries. Transblotting onto PVDF is carried out overnight according to standard procedures using a tris/glycine/methanol buffer system without SDS. The dried stained (see BP-2) or unstained blot is returned to the client. Staining facilitates matching to a duplicate Coomassie gel for mass spectrometry. Original gels are stained to make sure they’re blank and discarded, unless otherwise requested. We will change the blotting procedure to your specifications if necessary.
Price:

BP-1 (Standard Format): $70 per blot

BP-1LF (Large Format):  $80 per blot

BP-2: PVDF COOMASSIE STAINING.  The 2D gel pattern obtained from Coomassie blue staining of a PVDF membrane is faded but the scanned image (2D-ES-14, Electronic photos, $15) can be contrasted with Adobe photoshop. The image is super-imposable with the ECL film from Western blotting and helps to match a few spots on the film to a busy 2D gel pattern on a duplicate Coomassie gel run for mass spectrometry. Since Western blotting can be 100 times more sensitive than Coomassie, exact matching is very important. Coomassie staining of the blot does not interfere with subsequent Western blotting.
Price: add $12 per blot

BP-3: TRANSBLOTTING ONTO NITROCELLULOSE. Transblotting is carried out overnight according to standard gel procedures using a tris/glycine/methanol buffer system containing 0.1% SDS, with nitrocellulose of 0.45 micron pore size. Dried unstained blots are returned to the client and original gels are discarded unless additional treatments are requested. We will change the blotting procedure to your specifications if necessary.
Price:

BP-3 (Standard Format): $70 per blot

BP-3LF (Large Format): $80 per blot

BP-4: COLORED MOLECULAR WEIGHT MARKERS FOR BLOTS. These markers from Diversified Biotech are added during slab gel electrophoresis; so that colored bands are visible on both unstained gels and subsequent transblots at molecular weights of 95.5, 55, 43, 36, 29, 18.4, and 12.4 kDa. 
Price: $12 per gel.

BP-5: UBIQUITIN WESTERN BLOTTING PACKAGE INCLUDING ANTIBODY. We have optimized a method for identifying ubiquitin-containing proteins by 2D Western blotting using the Bethyl Labs anti-ubiquitin antibody (A300-317A). Our results suggest that the method is specific, reproducible, and sensitive. Note that this protein is very highly conserved so the antibody should react with all species.

Price:

BP-5SF (Standard Format Package): $325/sample includes standard format 2D gel electrophoresis, transblotting to PVDF, blot Coomassie staining, subsequent immunostaining with anti-ubiquitin antibody, and electronic photos of stained blot and ECL film.

BP-5SF-Dup (Replicate SF Package): $175 each

BP-5LF (Large Format Package): $400

BP-5LF-Dup (Replicate LF Package): $220 each

BP-6: WESTERN (IMMUNO-) BLOTTING, CLIENT SUPPLIES ANTIBODY.   This method for immuno-detection of specific bands or spots is carried out on PVDF transblots from 1-D or 2-D gels (see: "Handbook of Immunoblotting of Proteins Vol 1 and 11", Eds. Bjerrum, O. and Heegaar, N., 1988, CRC Press, Inc; and "Gel Electrophoresis: Proteins" by Dunn, M.J., 1993, Bios Scientific Publishers, Ltd). Requires prior electrophoresis (2D-ES-1 or 1D-ES-1) but PVDF transblotting (BP-1) is included in the fee. Generally the client supplies the antibody. The sensitive ECL method of detection is used. (Amersham reagents and protocol are used.) A control membrane run with secondary antibody is included as a control if necessary. 

BP-6SF Standard Format: Add $275/sample

BP-6SF-Dup: Add $100

BP-6LF  Large Format: Add $350/sample

BP-6LF-Dup: Add $140

BP-7: PHOSPHOTYROSINE WESTERN BLOTTING PACKAGE INCLUDING PY20 ANTIBODY. We have optimized a method for identifying phosphotyrosine-containing proteins by 2D Western blotting using the PY20 anti-PTyr antibody. Our results suggest that the method is specific, reproducible, and sensitive. However, in some cases the phosphate bond is labile so duplicate Western blots are encouraged to verify reproducibility. Mass spectrometry to identify proteins found by Western blotting is extra and requires a duplicate Coomassie blue stained gel. Ask for a quote by calling 800-462-3417 or emailing 2d@kendricklabs.com
Price:

BP-7SF (Standard Format): $325/sample includes standard format 2D gel electrophoresis, transblotting to PVDF, blot Coomassie staining, subsequent immunostaining with PY20 antibody, and electronic photos of stained blot and ECL film.

BP-7SF-Dup (Replicate SF Package): replicate W blot package $175 each

BP-7LF (Large Format Package): $400

BP-7LF-Dup (Replicate LF Package): $220 each

BP-8: COMBINED P-SERINE/P-THREONINE WESTERN BLOTTING PACKAGE USING ECL ADVANCE AND QIAGEN’S Q5 AND Q7 ABS. We have optimized a method for 2D Western blotting against phosphoserine and phosphothreonine residues on proteins using the Q5 and Q7 antibodies from Qiagen and ECL Advance from GE Healthcare. Both antibodies are advertised as detecting phosphorylated residues irrespective of surrounding amino acids. Note that non-specific binding shouldn't matter if control and test samples are compared for differences - it'd be the same in both. The ultra sensitive ECL Advance enables us to dilute the antibodies to 1:4000 and combine them for this package. The high sensitivity adds variability so all the BP-8 packages include duplicate Western blots to verify results.

BP-8SF (Standard Format): $575 includes duplicate SF 2D gel electrophoresis, duplicate transblotting to PVDF, blot Coomassie staining, subsequent immunostaining with combined Q5/Q7 antibodies, and electronic photos of stained blots and ECL films.

BP-8SF-Dup (Replicate SF Package): $200 each including 2D gel

BP-8LF (Large Format Package): $750 for package including duplicate blots

BP-8LF-Dup (Replicate LF Package): $275 each

BP-9: PHOSPHOSERINE OR PHOSPHOTHREONINE WESTERN BLOTTING. Same package as above except single antibodies are used instead of a mixture.
Price:

BP-9SF (Standard Format): $520 includes duplicate blots

BP-9SF-dup (SF Replicates): $180 each

BP-9LF (Large Format): $675 includes duplicate blots

BP-9LF-dup (LF Replicates): $250 each

BP-10: Acetyl-lysine Western Blotting. Acetylation is an important post translational modiciation occuring not only on histones but many other proteins.  2D electrophoresis and subsequent western bloting is a powerful tool for identifying acetylated proteins. Using Cell Signaling Technologies polyclonal acetylated lysine antibodies western blotting is carried out in a similar manner to the Ubiquitin western blotting package (BP-5).

BP-10SF (Standard Format) package: $325/sample includes standard format 2D gel electrophoresis, transblotting to PVDF, blot Coomassie staining, subsequent immunostaining with anti-acetylated lysine antibody, and electronic photos of stained blot and ECL film.

BP-10SF-Dup (SF Replicates): $175 each

BP-10LF (Large Format) package: $400 same package as BP10-SF except LF 2D gels are run.

BP-10LF (LF Duplicates): $220 each

Right: Acetylated proteins in rat liver. The anti-acetyl-lysine antibody from Cell Signaling was used for 2D gel WB at a 1:1000 dilution with overnight incubation. The  ECL film image from Western blotting is shown with permission of Blythe Shepard and Dr. Pamela Tuma, Catholic University of America, Washington, DC.  See also: Shepard, Blythe D., Dean J. Tuma, and Pamela L. Tuma. "Chronic Ethanol Consumption Induces Global Hepatic Protein Hyperacetylation." Alcohol Clin Exp Res 34.2 (2010): 1-12.

2D Gels/Top   Comparisons   Mass Spectrometry   Staining   Blotting   Radioactivity  Misc
 

SAMPLE PREPARATION PROCEDURES

SP-1: PROTEIN DETERMINATION followed by buffer addition. The total amount of protein in each sample is determined by the Pierce BCA total protein assay. If you require protein determinations, do not dissolve samples in buffers containing dithiothreitol or B-ME. We will add B-ME after protein analysis.
Price: $32 per sample.

SP-2 SPECIFIC ACTIVITY (PROTEIN-BOUND DPM) DETERMINATION This measurement includes TCA precipitation, washing the pellet and then determining cpm per mg protein. High concentrations of SDS (> 1.0%) add variability to this method. 
Price:  $32 per sample.

SP-3: ETHANOL PRECIPITATION OF PROTEINS followed by buffer addition. See the Ethanol Ppt Link
Price: $21 per sample.

SP-4: NUCLEASE TREATMENT  includes homogenization with osmotic lysis buffer containing protease inhibitors followed by incubation on ice with DNase and RNase to break down interfering polynucleotides. The composition of the 10X Nuclease Stock Solution is given in our Sample Preparation Guide. If you require nuclease treatment send the samples in a buffer containing less than 0.3% SDS. Higher SDS concentrations inactivate nucleases. 
Price:  $32 per sample.

SP-5: MICRODIALYSIS followed by lyophilization & buffer addition. High salt concentration (> 150 mM) interferes with IEF; the less salt the better.   If your samples contain high salt or buffer (phosphates, NaCl, etc.), this dialysis and concentration step is highly recommended.
Price: $25 per sample.

SP-6: LYOPHILIZATION followed by buffer addition.
Price: $22/sample.

SP-7: TCA PRECIPITATION followed by buffer addition. 
Price: $22/sample.

SP-8:  TCA/ACETONE PRECIPITATION followed by buffer addition. 
Price: $22/ sample.

SP-9: CUSTOM SAMPLE PREP. (Chunks of tissue, leaves, etc.) Charged by the hour.
Price: $150/ hr.

SP-11: ALBUMIN/IgG REMOVAL FROM SERUM, PLASMA & CSF OF HUMAN, RAT AND MOUSE using the Calbiochem ProteoExtract Albumin/IgG Removal kit (Cat. No. 122642) Depletion of albumin and IgG from 20-60 ul using the disposable gravity-flow affinity columns removes up to 75% of total serum proteins so that 3-4 times more enriched sample can be loaded on 2D gels.
Price: $50/sample.

2D Gels/Top   Comparisons   Mass Spectrometry   Staining  Blotting   Radioactivity   Sample Preparation 

cGMP-1 CURRENT GOOD MANUFACTURING PRACTICE The numerous SOPs required for cGMP for 1D and 2D electrophoresis are in place at Kendrick Labs along with an archival system for equipment calibrations and reports.

However, custom validation for the sample type of interest including SOPs are required for new projects to pass detailed FDA inspections - which may occur years down the road. Please let us know if you are going to submit an IND, or plan to use our  lab for QA of a product.
Price: Request Quote.

CON-1  CONSULTING FEES  Price: $180/hr. 

STORAGE, FREEZER LONG TERM: Kendrick Labs will store your samples 1 year in our –80° C freezer without charge. However, because of space constraints, we must now charge $150/6 months or $300/year for longer storage.
Price:

Storage-6M: $150

Storage-12M: $300