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Protocol - Fasting C-peptide - Assay

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Protocol Name from Source:

National Health and Nutrition Examination Survey (NHANES) Laboratory Procedures Manual

Availability:

Publicly available

Description:

This protocol provides instructions for drawing, processing, and storing blood according to the National Health and Nutrition Examination Survey (NHANES) methods. As there are no standard assays for C‑peptide, the protocol also provides basic guidelines to aid comparability among different studies.

Protocol:

The following is a summary version of the full National Health and Nutrition Examination Survey 2007-2008 protocol.

Exclusion Criteria

Persons will be excluded from this component if they:

• Report that they have hemophilia;

• Report that they have received cancer chemotherapy in the last 3 weeks; and

• Report that they have not fasted at least 9 hours.

SP = Sample Person.

1. Did you eat or drink anything other than plain water after [Insert time at 9 hours prior to sample collection] last night?

[ ] Yes

[ ] No

[ ] Refused

[ ] Don’t Know

If the answer is "No," then he or she has met the 9-hour fast. If the answer is "Yes," "Don’t know," or "Refused," then the actual fasting time is unknown.

Confirmation Question:

2. Have you had any of the following since {insert time from 1 here}?

Coffee or tea with cream and sugar? [Include milk or non-dairy creamers.]

[ ] Yes If Yes, record time and date _____________

[ ] No

Alcohol, such as beer, wine, or liquor?

[ ] Yes If Yes, record time and date _____________

[ ] No

Gum, breath mints, lozenges, or cough drops, or other cough or cold remedies?

[ ] Yes If Yes, record time and date _____________

[ ] No

Antacids, laxatives, or anti-diarrheals?

[ ] Yes If Yes, record time and date _____________

[ ] No

Dietary Supplements such as vitamins and minerals? [Include multivitamins and single nutrient supplements.]

[ ] Yes If Yes, record time and date _____________

[ ] No

3. Do you have hemophilia?

1 [ ] Yes

2 [ ] No

7 [ ] Refused

9 [ ] Don’t Know

If the SP answers "Yes," the SP is excluded from the blood draw. If the SP answers "No" or "Don’t Know," blood is drawn from the SP.

4. Have you received cancer chemotherapy in the past 4 weeks or do you anticipate such therapy in the next 4 weeks?

1 [ ] Yes

2 [ ] No

7 [ ] Refused

9 [ ] Don’t Know

If the SP answers "Yes," the SP is excluded from the blood draw. If the SP answers "No" or "Don’t Know," blood is drawn from the SP.

Venipuncture Procedures

Editor’s Note: Please review chapter 4 of the Laboratory Procedures Manual from the National Health and Nutrition Examination Survey for a full description of Phlebotomy procedures. [link[www.phenxtoolkit.org/toolkit_content/supplemental_info/diabetes/additional_info/NHANES_Lab_Manual.pdf|2007-2008 NHANES Lab Manual]].

Venipuncture should generally be performed using the median cubital, cephalic, or basilic veins in the left arm unless this arm is unsuitable. If the veins in the left arm are unsuitable, look for suitable veins on the right arm. If the veins in the antecubital space on both arms are not suitable, then look for veins in the forearm or dorsal side of the hand on the left arm/hand and then the right arm/hand.

Record the Results of the Venipuncture Procedure

Immediately after completing the venipuncture, record the results of the blood draw, the reasons for a tube not being drawn according to the protocol, and any comments about the venipuncture.

Process the Sample for the C-peptide Assay

Editor’s Note: Please review chapter 8 of the Laboratory Procedures Manual from the National Health and Nutrition Examination Survey 2007-2008 for a full description of Blood Processing procedures: [link[www.phenxtoolkit.org/toolkit_content/supplemental_info/diabetes/additional_info/NHANES_Lab_Manual.pdf|2007-2008 NHANES Lab Manual]].

• Allow the blood to clot by setting aside for 30-45 minutes at room temperature. Do not clot for more than 1 hour.

• Centrifuge the tube at room temperature to separate the serum and aliquot into an appropriate storage tube.

• Determine if the serum is hemolyzed, turbid, lipemic, or icteric. If so, enter a comment to describe the serum.

Laboratory Assay for C-peptide

The Diabetes Working Group (WG) notes that although there is not a standardized assay, there are a number of different kits that are appropriate to measure the concentration of C-peptide in serum. Once an assay is chosen for a particular study, the WG recommends that no changes in the protocol be made over the course of the study. To aid comparability, the WG recommends that Toolkit users follow manufacturer’s instructions and carefully record their procedure, including the make and manufacturer of equipment used and the repeatability and coefficients of variation for the assay.

Reference Range

Concentrations of fasting C-peptide normally range between 0.22 and 0.87 pmol/mL.

Personnel and Training Required

Phlebotomist Laboratory capable of performing c-peptide assay

Equipment Needs

Phlebotomy supplies

Requirements

Requirement CategoryRequired
Average time of greater than 15 minutes in an unaffected individualNo
Major equipmentNo
Specialized requirements for biospecimen collectionNo
Specialized trainingNo

Mode of Administration

Self-administered

Life Stage:

Child, Adolescent, Adult, Senior

Specific Instructions:

Although all three are valuable stand-alone measures and all should be collected if resources are available, the Diabetes Working Group (WG) recommends that investigators prioritize Insulin first, C‑peptide second, and Proinsulin third. This is because results from the Insulin bioassay can be combined with results from Fasting Plasma Glucose bioassay to derive the homeostatic model assessment (HOMA) of insulin resistance and pancreatic beta cell function. The Diabetes WG notes that while it is not the preferred method, the C-peptide assay can be performed on individuals who have not met the fasting requirements.

Note from the Diabetes WG: The investigator should record the reason a sample person is excluded from the blood draw, whether the subject is fasting, and the number of hours since the last meal.

Note from the Diabetes WG: Blood should be collected in an appropriate 5 mL or 10 mL red-top tube.

Note from the Diabetes WG: The Diabetes WG recommends that the investigator record whether the blood was drawn and whether the full amount was obtained.

Note from the Diabetes WG: Serum should be stored at -80°C until testing and shipped on dry ice to prevent thawing.

Note from the Expert Review Panel: Plasma or Serum

Collection of blood samples for the measurement of analytes requires a general determination of whether to use serum or plasma for the assay and also a determination of the type of collection tube to be obtained. For example, if serum is to be used, a determination needs to be made as to whether red-top or serum gel separator collection tubes are used. While comparable values are obtained for many analytes from either serum or plasma, there may be situations where differences are more pronounced, and serum- or plasma-specific norms will be needed for references. The protocol presented here uses red-top/serum separator tubes. At times, it may be possible to collect both, but other considerations, such as participant burden, may be the deciding factor. It is important to match assay type with sample type. Some automated devices may preclude the use of serum, for example, while others may be optimized for it. Investigators should choose methods of collection that match the methods of analysis. This will best be done by communicating with the laboratory where the proposed assays will be performed. The laboratory will become an important partner with you in assuring that there is compatibility from collection to assays to interpretation and reporting of levels and results.

Research Domain Information

Release Date:

October 1, 2015

Definition

A bioassay to measure serum concentration of C-peptide (connecting peptide), a small peptide that is cleaved from the proinsulin protein to form an active insulin protein. This peptide is produced by the pancreas as a by-product of insulin processing. It is secreted together with insulin.

Purpose

C-peptide levels may be used to differentiate type 1 from type 2 diabetes. People whose pancreas does not make any insulin (type 1 diabetes) have low levels of insulin and C-peptide. A person with type 2 diabetes has a normal or high level of C-peptide. C-peptide levels can be a more sensitive test of pancreas activity than insulin because they are not confounded by insulin replacement therapy. (Marques et al., 2004; ADA, 2010).

Selection Rationale

The National Health and Nutrition Examination Survey 2007-2008 protocol was selected as the best practice, standardized methodology for blood collection, processing, and storage.

Language

English, Spanish

Standards

StandardNameIDSource
Common Data Elements (CDE)Person Fasting C-peptide Measurement Value in pmol/ml3070892CDE Browser
Logical Observation Identifiers Names and Codes (LOINC)Fasting C-peptide proto62803-2LOINC

Process and Review

The [link[phenx.org/node/62|Expert Review Panel #1]] reviewed the measures in the Anthropometrics, Diabetes, Physical Activity and Physical Fitness, and Nutrition and Dietary Supplements domains.

Guidance from the ERP includes:

• Changed name of measure

• Added recommendations on use of serum or plasma

Back-compatible: no changes to Data Dictionary

Previous version in Toolkit archive ([link[www.phenxtoolkit.org/index.php?pageLink=browse.archive.protocols&id=140000|link]])

Source

Centers for Disease Control and Prevention (CDC), National Center for Health Statistics (NCHS). (2007). National Health and Nutrition Examination Survey Questionnaire. Laboratory Procedures Manual. Hyattsville, MD: U.S. Department of Health and Human Services, Centers for Disease Control and Prevention.

General References

American Diabetes Association (ADA). (2014). Diagnosis and classification of diabetes mellitus. Diabetes Care, 37(Suppl. 1), S81-S90.

Marques, R. G., Fontaine, M. J., & Rogers, J. (2004). C-peptide-Much more than a byproduct of insulin biosynthesis. Pancreas, 29(3), 231-238.

Miles, R. R., Roberts, R. F., Putnam, A. R., & Roberts, W. L. (2004). Comparison of serum and heparinized plasma samples for measurement of chemistry analytes [Letter to the Editor]. Clinical Chemistry, 50(9), 1704-1705.

Protocol ID:

141201

Variables:

Export Variables
Variable NameVariable IDVariable DescriptionVersiondbGaP Mapping
PX141201_Eat_Or_DrinkPX141201010000Did you eat or drink anything other than plain water after 11:30 last night?4N/A
PX141201_Last_Eat_Or_Drink_TimePX141201010100When did you last eat or drink anything other than plain?4Variable Mapping
PX141201_Last_Eat_Or_Drink_DatePX141201010200When did you last eat or drink anything other than plain water?4Variable Mapping
PX141201_Coffee_TeaPX141201020100Have you had any of the following since {insert time from 1 here}? Coffee or tea with cream and sugar? [Include milk or non-dairy creamers.]4Variable Mapping
PX141201_Coffee_Tea_TimePX141201020101If Yes, record time4N/A
PX141201_Coffee_Tea_DatePX141201020102If Yes, record Date4N/A
PX141201_AlcoholPX141201020201Have you had any of the following since {insert time from 1 here}? Alcohol, such as beer, wine, or liquor?4N/A
PX141201_Alcohol_TimePX141201020202If Yes, record time4N/A
PX141201_Alcohol_DatePX141201020203If Yes, record date4N/A
PX141201_GumPX141201020301Have you had any of the following since {insert time from 1 here}? Gum, breath mints, lozenges, or cough drops, or other cough or cold remedies?4N/A
PX141201_Gum_TimePX141201020302If Yes, record time4N/A
PX141201_Gum_DatePX141201020303If Yes, record date4N/A
PX141201_AntacidsPX141201020401Have you had any of the following since {insert time from 1 here}? Antacids, laxatives, or anti-diarrheals?4N/A
PX141201_Antacids_TimePX141201020402If Yes, record time4N/A
PX141201_Antacids_DatePX141201020403If Yes, record date4N/A
PX141201_Dietary_SupplementsPX141201020501Have you had any of the following since {insert time from 1 here}? Dietary Supplements such as vitamins and minerals? [Include multivitamins and single nutrient supplements.]4N/A
PX141201_Dietary_Supplements_TimePX141201020502If Yes, record time4N/A
PX141201_Dietary_Supplements_DatePX141201020503If Yes, record date4N/A
PX141201_HemophiliaPX141201030000Do you have hemophilia? (exclusion from Phlebotomy)4Variable Mapping
PX141201_ChemotherapyPX141201040000Have you received cancer chemotherapy in the past four weeks or do you anticipate such therapy in the next four weeks? (exclusion from Phlebotomy)4N/A
PX141201_Exclusion_CriteriaPX141201050000Exclusion Criteria4N/A
PX141201_Blood_Draw_DonePX141201060100Was blood drawn?4Variable Mapping
PX141201_Blood_Draw_SamplePX141201060200Was full sample obtained?4Variable Mapping
PX141201_Blood_Draw_CommentsPX141201060300Record any comments about the blood draw, including any reasons for the tube not being drawn according to the protocol.4Variable Mapping
PX141201_Sample_CommentsPX141201070000Comments to describe the sample during processing.4Variable Mapping
PX141201_Assay_TypePX141201080000Type of assay performed to determine the concentration of C-peptide.4N/A
PX141201_Equipment_Make_ManufacturerPX141201090000Make and manufacturer of the equipment used.4N/A
PX141201_Assay_RepeatabilityPX141201100000Repeatability of the assay4N/A
PX141201_Coefficient_Of_VariationPX141201110000Coefficient of variation for the assay4N/A
PX141201_C_Peptide_ConcentrationPX141201120000C-peptide concentration4N/A
PX141201_Fasting_Or_Casual_CPeptide_ConcentrationPX141201130000Was the participant fasting?4N/A