May 23, 2013
Sample Preparation and Volumes
Required per Assay
The Hormone Assay & Analytical Services Core of the Vanderbilt Diabetes Research & Training Center offers the following assays at the indicated cost per sample effected April 1, 2007. Please call and/or email Wanda Snead if you require additional information or clarification concerning prices or available assays.
| Assay | Requirements for Human and Canine Samples |
Requirements for Mouse Samples |
|
|---|---|---|---|
| 1 | Catecholamines (HPLC) | 250 microliters of plasma preserved in EGTA-glutathione for extraction 1-day assay. assay range: 0-2000 pg/ml | 50 micro liters plasma preserved in EGTA-glutathione. 1-day assay. assay range: 0-2000 pg/ml. |
| 2 | Insulin (RIA) |
225 micro liters of plasma for duplicate analysis. 2-day double Antibody Assay. assay range: 1 -150 micro units/ml. | 25 micro liters of plasma minimum for duplicate analysis. 5-day double antibody assay w/dilution procedure. Assay range: 0.1 - 2 ng/ml for 225 ul of sample (or 0.01-0.2 ng/ml for 25 ul of sample at dilution factor of 10). |
| 3 | "Free" Insulin (RIA) |
600 micro liters of plasma for peg-Extraction and duplicate analysis 2-day double antibody assay assay range: 1-150 micro units/ml | Routine Insulin assay (not "Free" for mouse) requires 225 micro liters of plasma. Perifusion media, or tissue culture, 2-day assay. Assay range: 0.1-10 ng/ml |
| 4 | Glucagon (RIA) |
225 micro liters of plasma for duplicate analysis preserved in 75 micro liters trasylol/ml of whole blood, 3-day double antibody assay: Assay range: 20-400 pg/ml | 25 micro liters plasma for duplicate analysis preserved in trasylol.5-day double antibody dilution based on dilution procedure. assay range: 5-200 pg/ml |
| 5 | Cortisol (RIA) |
60 micro liters of plasma for duplicate analysis .1-day coated tube assay Assay range: 0.5-50 dl/ml |
Corticosterone measurement in mice 15 micro liters of plasma for duplicate analysis 1-day double antibody procedure assay range: 12.5-1,000 ng/ml |
| 6 | Salivary Cortisol (RIA) |
400 micro liters for duplicate analysis 5-day double antibody Assay. Assay range :5-200ng/ml | N/A |
| 7 | Growth Hormone (RIA) |
225 micro liters for duplicate analysis.1-day IRMA(bead Assay) Assay range: 0.5 - 50 ng/ml | 25 micro liters for duplicate analysis 5-day double antibody procedure. Assay range: 0.2- 2 ng/ml for 225 ul of sample (or 2-20 ng/ml for 25 ul of sample) |
| 8 | Leptin (RIA) |
225 micro liters for duplicate analysis.2-day double antibody assay. Assay Range : 0.5-100 ng/ml. | 25 micro liters of plasma for duplicate analysis. 5-day double antibody procedure < AssayRange : 0.1- 2 ng/ml for 225 ul of sample (or 1-20 ng/ml for 25 ul of sample) |
| 9 | Pancreatic Polypeptide (RIA) |
225 micro liters for duplicate analysis.3-day double antibody assay. Trasylol is also added. Assay range: 20-1200 pg/ml | N/A |
| 10 | C-peptide (RIA) |
225 micro liters for duplicate analysis in trasylol.1-3 day double antibody assay depending on species Assay range: 0.1-10ng/ml |
25 micro liters of plasma for duplicate analysis. Involves a dilution step and is a 5-day double antibody procedure. Assay Range: 0.016- 1.6 ng/ml for 225 ul of sample (or 0.16-16 ng/ml for 25 ul of sample) |
| 11 | IGF-1 (RIA) |
300 micro liters for duplicate analysis. 5-day double antibody assay. Assay Range: | 30 micro liters of plasma for extraction and duplicate analysis. 5-day double antibody procedure. Assay Range: |
| 12 | ACTH (RIA) |
ACTH (RIA) 225 micro liters for duplicate analysis.5-day double antibody assay. Assay Range : 4-150 pg/ml | 55 micro liters of plasma for duplicate analysis. Involves a dilution step and is a 5-day double antibody procedure. Assay Range: 4- 150 pg/ml for 225 ul of sample (or 16-600 pg/ml for 55 ul of sample). |
| 13 | Aldosterone (RIA) |
225 micro liters for duplicate analysis. 2-day double antibody assay. Assay Range: 2-200 pg/ml | 45 micro liters of plasma for duplicate analysis. Involves a dilution step and is a 2-day double antibody procedure. Assay Range: 2- 200 pg/ml for 225 ul of sample (or 10-1,000 pg/ml for 45 ul of sample). |
| 14 | Prolactin (RIA) |
N/A | 25 micro liters of plasma for dilution. 5- Day double antibody assay. Assay range: 1.5 - 20 ng/ml. |
| 15 | TSH (RIA) |
N/A | 55 micro liters of plasma for duplicate analysis. Involves a dilution step and is a 5-day double antibody procedure. Assay Range: 0.1- 2 ng/ml for 225 ul of sample (or 0.4-8 ng/ml for 55 ul of sample). |
| 16 | T3/T4 (RIA) |
N/A | 25 micro liters of plasma for dilution. 5-day double antibody assay. |
| 17 | Testosterone (RIA) |
25 micro liters of plasma for dilution and duplicate analysis in a 1-day double antibody assay. Assay Range: 0.06-15.6 ng/ml | 25 micro liters of plasma for dilution and duplicate analysis in a 1-day double antibody assay. Assay Range: 0.06-15.6 ng/ml |
| 18 | Estradiol (RIA) |
25 micro liters of plasma for dilution and duplicate analysis in a 1-day double antibody assay. Assay Range: 8-240 pg/ml | 25 micro liters of plasma for dilution and duplicate analysis in a 1-day double antibody assay. Assay Range: 8-240 pg/ml |
| 19 | Full Profile and
Select Amino Acids (HPLC) |
500 micro liters of plasma, or call for whole blood and low sample volume prep | 50 micro liters of plasma, or call for whole blood and low sample volume prep |
| 20 | Adiponectin & Resistin | 15 micro liters for duplicate analysis, multiplexed luminex assay | 15 micro liters for duplicate analysis, multiplexed luminex assay |
| 21 | Adiponectin & Resistin | N/A | 25 micro liters of plasma into 500 micro liters of cold acetonitrile, dry, reconstitute in 200 micro liters |
| 22 | UDGP/PEP | 500mg tissue labeled with 3H or 14C in liquid N2 (HPLC) | N/A |
| 23 | cAMP | 500mg of insitu collected liver homogenized using 1 ml ice cold .4M PCA containing .5mM EGTA. After 1 min on ice, centrifuge at ~3500g for 10 min at 4 C. The supernatant neutralized with 360ul .5M K@CO3 pH 6.8. The neutralized solution is spun again for 5 min to ppt insoluable KClO4. HPLC | N/A |
| 24 | Purine Nucleotides | Insitu collected and weighed, 100mg of liver (liquid nitrogen) add 1ml cold .4M perchloric acid with .5M EGTA.homogenize, sit 10minutes ice, spin cold ~3200g 10minutesdecant supernatant-add 8drops .5M K2CO3, mix, ice 5minutes spin cold ~3200g 5 minutes, decant; save supernatant, freeze/ analyze.(HPLC) | Insitu collected and weighed, 100mg of liver (liquid nitrogen) add 1ml cold .4M perchloric acid with .5M EGTAhomogenize, sit 10minutes ice, spin cold ~3200g 10minutesdecant supernatant-add 8 drops .5M K2CO3, mix, ice 5minutes spin cold ~3200g 5minutes, decant; save supernatant, freeze, analyze (HPLC) |
| 25 | Specific Activity (Alanine, leucine & phenylanine) |
750 micro liters of plasma Separated individually with heated cation exchange columns, sodium citrate buffers and proportioning pump for splitting the column effluent to a fraction collector for scintillation counting, with the remaining flow being mixed with ninhydrin, heated before entering a colorimeter for determining concentration. (HPLC) | N/A |
| 26 | 8-isoprostane-GF2a (also referred to as F2-IsoPs) |
Plasma, 3mL. Collect in tubes containing EDTA and store immediately at -80oC, do not thaw. Urine, 1mL, Store at -80oC after collection. Tissues, 100mg, flash freeze in liquid nitrogen immediately, do not thaw.CSF, 1mL, Store at -80oC after collection. | N/A |
| 27 | 8-isoprostane-PGF2a | metabolite (urine, 1mL, store at -80oC). | N/A |
| 28 | Isofurans | urine, tissues, CSF, same requirements as IsoP analysis | N/A |
| 29 | Neuroprostanes | Tissues, same requirements as IsoP analysis. Derived from the oxidation of docosahexaenoic acid (DHA). DHA is abundant in the brain and is found in fish oil. All urinary assays require an additional 0.5mL urine for creatinine. | N/A |
| 30 | Prostaglandin Analysis: | Prostaglandins (PGs) F2a, D2, E2, J2, A2 (tissues, 100mg).PGE2 metabolite (urine, 1mL).PGD2 metabolite (urine, 3mL)Prostacyclin PGI2) metabolite (2,3-dinor-6-keto-PGF1a) (urine, 3mL).Thromboxane metabolite (11-dehydro-thromboxane B2) (urine, 3mL). Leukotriene E4 (urine, 5mL) | Special Handling: Tissues hould be flash frozen in liquid nitrogen and stored at -80oC. All urine should be stored at -80oC after collection. |
| 31 | Total cholesterol | 10 ul serum | 10 ul serum |
| 32 | Triglycerides | 10 ul serum | 10 ul serum |
| 33 | HDL cholesterol | 30 ul serum | 30 ul serum |
The Vanderbilt Diabetes Research and Training Center is supported by NIH grant DK20593.
