www.caymanchem.comCustomer Service 800.364.9897Technical Support 888.526.53511180 E. Ellsworth Rd · Ann Arbor, MI · USA

Albumin (human) ELISA Kit

Item No. 501400

3GENERAL INFORMATION

TABLE OF CONTENTS GENERAL INFORMATION 3 Materials Supplied

4 Safety Data4 Precautions4 If You Have Problems5 Storage and Stability5 Materials Needed but Not Supplied

INTRODUCTION 6 Background6 About This Assay7 Principle of the Assay9 Definition of Key Terms

PRE-ASSAY PREPARATION 10 Buffer Preparation11 Sample Preparation

ASSAY PROTOCOL 13 Preparation of Assay-Specific Reagents15 Plate Set Up16 Performing the Assay

ANALYSIS 19 Calculations21 Performance Characteristics

RESOURCES 24 Troubleshooting25 References26 Plate Template27 Notes27 Warranty and Limitation of Remedy

GENERAL INFORMATION

Materials Supplied

Item Number

Item 96 wells Quantity/Size

401402 Albumin (human) ELISA Monoclonal Antibody 1 vial/100 dtn

401400 Albumin (human) AP Tracer 1 vial/100 dtn

401404 Albumin (human) ELISA Standard 1 vial

400080 Tris Buffer Concentrate (10X) 2 vials/10 ml

411007 AP Wash Buffer Concentrate (150X) 1 vial/5 ml

400009 Precoated (Goat Anti-Mouse IgG) ELISA 96-Well Strip Plate 1 plate

400012 96-Well Cover Sheet 1 cover

400090 pNPP Tablets 1 vial/5 ea

400082 DEA Buffer Concentrate (10X) 1 vial/2.5 ml

If any of the items listed above are damaged or missing, please contact our Customer Service department at (800) 364-9897 or (734) 971-3335. We cannot accept any returns without prior authorization.

! WARNING: THIS PRODUCT IS FOR RESEARCH ONLY - NOT FORHUMAN OR VETERINARY DIAGNOSTIC OR THERAPEUTIC USE.

4 GENERAL INFORMATION 5GENERAL INFORMATION

Safety DataThis material should be considered hazardous until further information becomes available. Do not ingest, inhale, get in eyes, on skin, or on clothing. Wash thoroughly after handling. Before use, the user must review the complete Safety Data Sheet, which has been sent via email to your institution.

PrecautionsPlease read these instructions carefully before beginning this assay.This kit may not perform as described if any reagent or procedure is replaced or modified.When compared to quantification by LC/MS or GC/MS, it is not uncommon for immunoassays to report higher analyte concentrations. While LC/MS or GC/MS analyses typically measure only a single compound, antibodies used in immunoassays sometimes recognize not only the target molecule, but also structurally related molecules, including biologically relevant metabolites. In many cases, measurement of both the parent molecule and metabolites is more representative of the overall biological response than is the measurement of a short-lived parent molecule. It is the responsibility of the researcher to understand the limits of both assay systems and to interpret their data accordingly.

If You Have ProblemsTechnical Service Contact Information

Phone: 888-526-5351 (USA and Canada only) or 734-975-3888Fax: 734-971-3641Email: techserv@caymanchem.comHours: M-F 8:00 AM to 5:30 PM EST

In order for our staff to assist you quickly and efficiently, please be ready to supply the lot number of the kit (found on the outside of the box).

Storage and StabilityThis kit will perform as specified if stored as directed at -20°C and used before the expiration date indicated on the outside of the box.

Materials Needed But Not Supplied1. A plate reader capable of measuring absorbance at 405 nm.2. Adjustable pipettes and a repeating pipettor.3. A source of ‘UltraPure’ water. Water used to prepare all ELISA reagents and

buffers must be deionized and free of trace organic contaminants (‘UltraPure’). Use activated carbon filter cartridges or other organic scavengers. Glass distilled water (even if double distilled), HPLC-grade water, and sterile water (for injections) are not adequate for ELISA. NOTE: UltraPure water is available for purchase from Cayman (Item No. 400000).

4. Materials used for Sample Preparation (see page 11).

6 INTRODUCTION 7INTRODUCTION

INTRODUCTION

BackgroundAlbumin makes up approximately 50% of the protein in human serum. Albumin has many important physiological functions including maintaining osmotic pressure, buffering pH of the blood, and the transport of hormones, fatty acids, and other molecules throughout the body.The measurement of albumin in urine is well established clinically as an indicator of renal function. Elevated levels of albumin (microalbuminuria and macroalbuminuria) in the urine are an indication of various disease states including diabetic nephropathy, non-diabetic kidney disease, and cardiovascular morbidity and mortality.1-3 Normal urinary albumin levels of adults are less than 30 µg albumin per mg of creatinine. Microalbuminuria is commonly defined as 30-300 µg albumin per mg of creatinine and macroalbuminuria is commonly defined as albumin levels greater than 300 µg/mg.4,5

About This AssayCayman’s Albumin (human) ELISA Kit is a competitive assay that can be used for quantification of human albumin in urine. The assay has a range from 15.6-2,000 ng/ml and a sensitivity (80% B/B0) of approximately 60 ng/ml.

Principle of the AssayThis assay is based on the competition between albumin and an albumin-alkaline phosphatase (AP) conjugate (Albumin Tracer) for a limited amount of Albumin Monoclonal Antibody. Because the concentration of the Albumin Tracer is held constant while the concentration of albumin varies, the amount of Albumin Tracer that is able to bind to the Albumin Monoclonal Antibody will be inversely proportional to the concentration of albumin in the well. This antibody-albumin complex binds to goat anti-mouse IgG that has been previously attached to the well. The plate is washed to remove any unbound reagents and then pNPP is added to the well. The product of this enzymatic reaction has a distinct yellow color and absorbs strongly at 405 nm. The intensity of this color, determined spectrophotometrically, is proportional to the amount of Albumin Tracer bound to the well, which is inversely proportional to the amount of free albumin present in the well during the incubation; or

Absorbance ∝ [Bound Albumin Tracer] ∝ 1/[Albumin]A schematic of this process is shown in Figure 1, see page 8.

8 INTRODUCTION 9INTRODUCTION

Definition of Key Terms

Blank: background absorbance caused by pNPP Reagent. The blank absorbance should be subtracted from the absorbance readings of all the other wells, including NSB wells.

Total Activity: total enzymatic activity of the AP-linked tracer. This is analogous to the specific activity of a radioactive tracer.

NSB (Non-Specific Binding): non-immunological binding of the tracer to the well. Even in the absence of specific antibody a very small amount of tracer still binds to the well; the NSB is a measure of this low binding. Do not forget to subtract the Blank absorbance values.

B0 (Maximum Binding): maximum amount of the tracer that the antibody can bind in the absence of free analyte.

%B/B0 (%Bound/Maximum Bound): ratio of the absorbance of a particular sample or standard well to that of the maximum binding (B0) well.

Standard Curve: a plot of the %B/B0 values versus concentration of a series of wells containing various known amounts of analyte.

Dtn: determination, where one dtn is the amount of reagent used per well.

1. Incubate with tracer, an�body, and either standard or sample.

2. Wash to remove all unbound reagents.

3. Develop the well with pNPP solu�on.

Wells are pre-coated with goat an�-mouse IgG and blocked with a proprietary formula�on of proteins.

= Goat An�-Mouse IgG

= Blocking proteins

= Alkaline Phospharase linked

to Albumin (tracer)

= Specific an�body to Albumin (human)

= Free Albumin

Figure 1. Schematic of the ELISA

10 PRE-ASSAY PREPARATION 11PRE-ASSAY PREPARATION

PRE-ASSAY PREPARATION

NOTE: Water used to prepare all ELISA reagents and buffers must be deionized and free of trace organic contaminants (‘UltraPure’). Use activated carbon filter cartridges or other organic scavengers. Glass distilled water (even if double distilled), HPLC-grade water, and sterile water (for injections) are not adequate for ELISA. UltraPure water may be purchased from Cayman (Item No. 400000).

Buffer PreparationStore all diluted buffers at 4°C; they should be stable for about two months.1. Tris Buffer Preparation

Dilute the contents of one vial of Tris Buffer Concentrate (10X) (Item No. 400080) with 90 ml of UltraPure water. Be certain to rinse the vial to remove any salts that may have precipitated. NOTE: It is normal for the concentrated buffer to contain crystalline salts after thawing. These will completely dissolve upon dilution with water.

2. DEA Buffer PreparationDilute the contents of one vial of DEA Buffer Concentrate (10X) (Item No. 400082) to a total volume of 25 ml with UltraPure water.

3. AP Wash Buffer PreparationDilute the contents of one vial of AP Wash Buffer Concentrate (150X) (Item No. 411007) to a total volume of 750 ml with UltraPure water. NOTE: Smaller volumes of AP Wash Buffer can be prepared by diluting the concentrate 1:150.

Sample PreparationThis assay has been validated for the quantification of albumin in human urine (see Figure 2, on page 12). Proper sample storage and preparation are essential for consistent and accurate results. Please read this section thoroughly before beginning the assay.

General Precautions • All samples must be free of organic solvents prior to assay.• Samples should be assayed immediately after collection; samples that

cannot be assayed immediately should be stored at -80°C.

UrineSamples should be centrifuged and tested immediately after collection. If samples cannot be assayed immediately, they should be centrifuged and frozen at -80°C.3

Since most urine samples will need to be diluted at least 1:4 (i.e., 250 µl into 1 ml of buffer) to fall within the standard curve of this assay, interference should not be a problem. To determine the proper dilution range for your samples, dilute one or two test samples in Tris Buffer to obtain at least two different dilutions of each sample between 771 and 64 ng/ml (i.e., between 20-80% B/B0).NOTE: Samples should be standardized to creatinine levels in the urine. Creatinine (urinary) Colorimetric Assay Kit (Item No. 500701) is a quick, convenient method for measuring creatinine in urine.

13ASSAY PROTOCOL12 PRE-ASSAY PREPARATION

40

60

80

100

Alb

um

in m

easu

red

in s

amp

le (

µg

/ml)

Concentration of albumin added to urine sample (µg/ml)

0 20 40 600

80

20

Slope = 1.035y-intercept = 5.24 µg/mlr2 = 0.995

Figure 2. Recovery of albumin from urine. Urine samples were spiked with albumin, diluted as described in the Sample Preparation section and analyzed using the Albumin (human) ELISA Kit. The y-intercept corresponds to the amount of albumin in unspiked urine. Error bars repesent standard deviations obtained from multiple diutions of each sample.

ASSAY PROTOCOL

Preparation of Assay-Specific ReagentsAlbumin (human) ELISA StandardThis material should be considered a potential biohazard until information to the contrary becomes available. Do not ingest, swallow, or inhale. Do not get in eyes, on skin, or on clothing. Wash thoroughly after handling. This information contains some, but not all, of the information required for the safe and proper use of this material. Before use, the user must review the complete Material Safety Data Sheet, which has been sent via email to your institution. Reconstitute the contents of the Albumin (human) ELISA Standard (Item No. 401404) with 1 ml of Tris Buffer. The concentration of this solution (the bulk standard) will be 20 µg/ml. Stored at 4°C; this standard should be stable for up to four weeks.To prepare the standard for use in ELISA: Obtain eight clean test tubes and number them #1 through #8. Aliquot 900 µl Tris Buffer to tube #1 and 500 µl Tris Buffer to tubes #2-8. Transfer 100 µl of the bulk standard (20 µg/ml) to tube #1 and mix thoroughly. The concentration of this standard, the first point on the standard curve, will be 2,000 ng/ml. Serially dilute the standard by removing 500 µl from tube #1 and placing in tube #2; mix thoroughly. Next, remove 500 µl from tube #2 and place it into tube #3; mix thoroughly. Repeat this process for tubes #4-8. These diluted standards should not be stored for more than 24 hours.

14 ASSAY PROTOCOL 15ASSAY PROTOCOL

20 µg/mlBulk Standard

100 µl 500 µl 500 µl 500 µl 500 µl 500 µl 500 µl 500 µl

900 µlTris

Bu�er

500 µlTris

Bu�er

Final

2,000ng/ml

S1 S2 S3 S4 S5 S6 S7 S8

1,000ng/ml

500ng/ml

250ng/ml

125ng/ml

62.5ng/ml

31.3ng/ml

15.6ng/ml

500 µlTris

Bu�er

500 µlTris

Bu�er

500 µlTris

Bu�er

500 µlTris

Bu�er

500 µlTris

Bu�er

500 µlTris

Bu�er

1 ml Tris Bu�er

Figure 3. Preparation of the albumin standards

Albumin (human) AP TracerReconstitute the Albumin (human) AP Tracer (Item No. 401400) with 6 ml Tris Buffer. Store the reconstituted tracer at 4°C (do no freeze!) and use within four weeks. A 20% surplus of tracer has been included to account for any incidental losses.Albumin (human) ELISA Monoclonal AntibodyReconstitute the Albumin (human) ELISA Monoclonal Antibody (Item No. 401402) with 6 ml Tris Buffer. Store the reconstituted antibody at 4°C and use within four weeks. A 20% surplus of antibody has been included to account for any incidental losses.

Plate Set UpThe 96-well plate(s) included with this kit is supplied ready to use. It is not necessary to rinse the plate(s) prior to adding the reagents. NOTE: If you do not need to use all the strips at once, place the unused strips back in the plate packet and store at 4°C. Be sure the packet is sealed with the desiccant inside. Each plate or set of strips must contain a minimum of two blanks (Blk), two non-specific binding wells (NSB), two maximum binding wells (B0), and an eight point standard curve run in duplicate. NOTE: Each assay must contain this minimum configuration in order to ensure accurate and reproducible results. Each sample should be assayed at two dilutions and each dilution should be assayed in duplicate. For statistical purposes, we recommend assaying samples in triplicate.A suggested plate format is shown in Figure 4, below. The user may vary the location and type of wells present as necessary for each particular experiment. The plate format provided below has been designed to allow for easy data analysis using a convenient spreadsheet offered by Cayman (see page 19, for more details). We suggest you record the contents of each well on the template sheet provided (see page 26).

Blk - BlankTA - Total Ac�vityNSB - Non-Specific BindingB0 - Maximum BindingS1-S8 - Standards 1-81-24 - Samples

A

B

C

D

E

F

G

H

1 2 3 4 5 6 7 8 9 10 11 12S1

S2

S3

S4

S5

S6

S7

S8 S8

S7

S6

S5

S4

S3

S2

S1

8

7

6

5

4

3

2

1

8

7

6

5

4

3

2

1

8

7

6

5

4

3

2

1

16

15

14

13

12

11

10

9

16

15

14

13

12

11

10

9

16

15

14

13

12

11

10

9

24

23

22

21

20

19

18

17

24

23

22

21

20

19

18

17 17

24

23

22

21

20

19

18

Blk

Blk

NSB

NSB

B0

B0

B0

TA

Figure 4. Sample plate format

16 ASSAY PROTOCOL 17ASSAY PROTOCOL

Performing the Assay

Pipetting Hints

• Use different tips to pipette each reagent.• Before pipetting each reagent, equilibrate the pipette tip in that

reagent (i.e., slowly fill the tip and gently expel the contents, repeat several times).

• Do not expose the pipette tip to the reagent(s) already in the well.

Addition of the Reagents1. Tris Buffer

Add 100 µl Tris Buffer to NSB wells. Add 50 µl Tris Buffer to B0 wells. 2. Albumin (human) ELISA Standard

Add 50 µl from tube #8 to both of the lowest standard wells (S8). Add 50 µl from tube #7 to each of the next two standard wells (S7). Continue with this procedure until all the standards are aliquoted. The same pipette tip should be used to aliquot all the standards. Before pipetting each standard, be sure to equilibrate the pipette tip in that standard.

3. SamplesAdd 50 µl of sample per well. Each sample should be assayed at a minimum of two dilutions. Each dilution should be assayed in duplicate (triplicate recommended).

4. Albumin (human) AP TracerAdd 50 µl to each well except the TA and the Blk wells.

5. Albumin (human) ELISA Monoclonal AntibodyAdd 50 µl to each well except the TA, the NSB, and the Blk wells.

Well Tris Buffer Standard/Sample Tracer Antibody

Blk - - - -

TA - - 5 µl (at devl. step) -

NSB 100 µl - 50 µl -

B0 50 µl - 50 µl 50 µl

Std/Sample - 50 µl 50 µl 50 µl

Table 1. Pipetting summary

Incubation of the PlateCover each plate with plastic film (Item No. 400012) and incubate for two hours at room temperature on an orbital shaker.

19ANALYSIS18 ASSAY PROTOCOL

Development of the Plate1. When ready to develop the plate, reconstitute 5 pNPP tablets in 25 ml DEA

Buffer (25 ml is sufficient to develop a 96-well plate). Reconstituted pNPP is not stable and we recommend that you store the reconstituted pNPP for no more than 24 hours. If the entire plate is not used at once, dissolve just enough pNPP for the number of wells used (1 pNPP tablet per 5 ml DEA Buffer).

2. Empty the wells and rinse four times with Wash Buffer. Each well should be completely filled with Wash Buffer during each wash. Invert the plate between wash steps to empty the fluid from the wells. After the last wash, gently tap the inverted plate on lint free absorbent paper to remove the residual Wash Buffer.

3. Add 200 µl of pNPP Substrate Solution to each well of the plate.4. Add 5 µl of tracer to the TA wells.5. Cover the plate with plastic film. Optimum development is obtained by using

an orbital shaker equipped with a large, flat cover to allow the plate(s) to develop in the dark. Incubate shaking for 60 minutes.

Reading the Plate1. Wipe the bottom of the plate with a lint free towel to remove fingerprints,

dirt, etc. 2. Read the plate at a wavelength of 405 nm.

ANALYSISMany plate readers come with data reduction software that plot data automatically. Alternatively a spreadsheet program can be used. The data should be plotted as either %B/B0 versus log concentration using a four-parameter logistic fit or as logit B/B0 versus log concentration using a linear fit. NOTE: Cayman has a computer spreadsheet available for data analysis. Please contact Technical Service or visit our website (www.caymanchem.com/analysis/elisa) to obtain a free copy of this convenient data analysis tool.

Calculations

Preparation of the DataThe following procedure is recommended for preparation of the data prior to graphical analysis.NOTE: If the plate reader has not subtracted the absorbance readings of the blank wells from the absorbance readings of the rest of the plate, be sure to do that now.1. Average the absorbance readings from the NSB wells.2. Average the absorbance readings from the B0 wells.3. Subtract the NSB average from the B0 average. This is the corrected B0 or

corrected maximum binding.4. Calculate the B/B0 (Sample or Standard Bound/Maximum Bound) for the

remaining wells. To do this, subtract the average NSB absorbance from the S1 absorbance and divide by the corrected B0 (from Step 3). Repeat for S2-S8 and all sample wells. (To obtain %B/B0 for a logistic four-parameter fit, multiply these values by 100.)

NOTE: The TA values are not used in the standard curve calculations. Rather, they are used as a diagnostic tool; the corrected B0 divided by the actual TA (10X measured absorbance) will give the %Bound. This value should closely approximate the %Bound that can be calculated from the Sample Data (see page 21). Erratic absorbance values and a low (or no) %Bound could indicate the presence of organic solvents in the buffer or other technical problems (see page 24 for Troubleshooting).

20 ANALYSIS 21ANALYSIS

Plot the Standard CurvePlot %B/B0 for standards S1-S8 versus albumin concentration using linear (y) and log (x) axes and perform a 4-parameter logistic fit.Alternative Plot - The data can also be lineraized using a logit transformation. The equation for this conversion is shown below. NOTE: Do not use %B/B0 in this calculation.

logit (B/B0) = ln [B/B0/(1 - B/B0)]

Plot the data as logit (B/B0) versus log concentrations and perform a linear regression fit.

Determine the Sample ConcentrationCalculate the B/B0 (or %B/B0) value for each sample. Determine the concentration of each sample using the equation obtained from the standard curve plot. NOTE: Remember to account for any concentration or dilution of the sample prior to the addition to the well. Samples with %B/B0 values greater than 80% or less than 20% should be re-assayed as they generally fall out of the linear range of the standard curve.

Performance Characteristics

Sample DataThe standard curve presented here is an example of the data typically produced with this kit; however, your results will not be identical to these. You must run a new standard curve. Do not use the data below to determine the value of your samples. Your results could differ substantially. Raw Data Average CorrectedTotal Activity 0.490 0.487 0.489NSB 0.006 0.001 0.003B0 1.099 1.034 1.173 0.958 1.066 1.063

Dose (ng/ml) Raw Data Corrected %B/B0

2,000 0.113 0.106 0.110 0.103 9.7 9.6

1,000 0.201 0.202 0.198 0.199 17.4 18.5

500 0.349 0.324 0.346 0.321 30.3 29.8

250 0.552 0.508 0.549 0.505 48.1 46.9

125 0.737 0.700 0.734 0.697 64.3 64.8

62.5 0.900 0.883 0.897 0.880 78.6 81.8

31.3 0.983 0.974 0.980 0.971 85.9 90.2

15.6 1.099 1.034 1.096 1.031 96.0 95.9

Table 2. Typical results

22 ANALYSIS 23ANALYSIS

0

20

40

60

80

100

0

20

40

60

80

100

Albumin (human) (ng/ml)

%B

/B0 __

__

%C

V ---

-

10 100 1,000 10,000

Assay Range = 15.6-2,000 ng/mlSensitivity (defined as 80% B/B0) = 60 ng/mlMid-point (defined as 50% B/B0) = 215 ng/ml

The sensitivity and mid-point were derived from the standard curve shown above. The standard was diluted with Tris Buffer.

Evaluate data cautiously

Use data with confidence

Figure 5. Typical standard curve

Precision:The intra- and inter-assay CVs have been determined at multiple points on the standard curve. These data are summarized in the graph on page 22 and in the table below.

Dose (ng/ml) %CV* Intra-assay variation

%CV* Inter-assay variation

2,000 11.2 6.5

1,000 7.4 4.9

500 12.0 3.3

250 11.0 4.4

125 14.8 4.6

62.5 27.4 6.6

31.3 38.6 13.5

15.6 55.2 32.7

Table 3. Intra- and inter-assay variation*%CV represents the variation in concentration (not absorbance) as determined using a reference standard curve.

Cross Reactivity:This assay has been validated for measuring albumin in human urine samples. Both mouse and rat albumins have been tested in this assay and found to be completely non-cross reactive.

24 RESOURCES 25RESOURCES

RESOURCES

Troubleshooting

Problem Possible Causes Recommended Solutions

Erratic values; dispersion of duplicates

A. Trace organic contaminants in the water source

B. Poor pipetting/technique

A. Replace activated carbon filter or change source of UltraPure water

High NSB (>10% of B0) A. Poor washing B. Exposure of NSB wells to

specific antibody

A. Rewash plate and redevelop

Very low B0 A. Trace organic contaminants in the water source

B. Plate requires additional development time

C. Dilution error in preparing reagents

A. Replace activated carbon filter or change source of UltraPure water

B. Return plate to shaker and re-read later

Low sensitivity (shift in dose response curve)

Standard is degraded Replace standard

Analyses of two dilutions of a biological sample do not agree (i.e., more than 20% difference)

Interfering substances are present

Purify sample prior to analysis by ELISA6

Only Total Activity (TA) wells develop

Trace organic contaminants in the water source

Replace activated carbon filter or change source of UltraPure water

References1. Hovind, P., Tarnow, L., Rossing, P., et al. Predictors for the development of

microalbuminuria and macroalbuminuria in patients with type 1 diabetes: Inception cohort study. BMJ 328, 1105-1110 (2004).

2. Damsgaard, E.M., Froland, A., Jorgensen, O.D., et al. Microalbuminuria as predictor of increased mortality in elderly people. Br. Med. J. 300, 297-300 (1990).

3. Brinkman, J.W., de Zeeuw, D., Duker, J.J., et al. Falsely low urinary albumin concentrations after prolonged frozen storage of urine samples. Clin. Chem. 51(11), 2181-2183 (2005).

4. Justesen, T.I., Petersen, J.L.A., Ekbom, P., et al. Albumin-to-creatinine ratio in random urine samples might replace 24-h urine collections in screening for micro- and macroalbuminuria in pregnant woman with type 1 diabetes. Diabetes Care 29(4), 924-925 (2006).

5. Ruggenenti, P., Gaspari, F., Perna, A., et al. Cross sectional longitudinal study of spot morning urine protein: Creatinine ratio, 24 hour urine protein excretion rate, glomerular filtration rate, and end stage renal failure in chronic renal disease in patients without diabetes. BMJ 316, 504-509 (1998).

6. Maxey, K.M., Maddipati, K.R. and Birkmeier, J. Interference in enzyme immunoassays. J. Clin. Immunoassay 15, 116-120 (1992).

26 RESOURCES 27RESOURCES

A B C D E F G H

12

34

56

78

910

1112

NOTES

Warranty and Limitation of RemedyBuyer agrees to purchase the material subject to Cayman’s Terms and Conditions.Complete Terms and Conditions including Warranty and Limitation of Liability information can be found on our website.This document is copyrighted. All rights are reserved. This document may not, in whole or part, be copied, photocopied, reproduced, translated, or reduced to any electronic medium or machine-readable form without prior consent, in writing, from Cayman Chemical Company.©07/13/2016, Cayman Chemical Company, Ann Arbor, MI, All rights reserved. Printed in U.S.A.