rev:April 17, 1997

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Cytokines, Growth Factors & Chemokines: BACKGROUND INFORMATION

-click for additional background on chemokines

-Originating from a single fertilized egg-cell, the mature human body contains some 85 trillion cells. Each of these cells has its own needs, as well as distinct duties, mots of which requires the collaboration with other cells. For the dynamic system of the body to be in perfect health, this huge assembly of individual cells must act collectively and in total harmony. This necessitates an efficient and reliable system for intercellular communication. The fundamental elements of this complex communication network are cytokines and their cellular receptors. Cytokines are soluble proteins produced and released by individual cells for the purpose of transmitting distinct messages of activation, inhibition, chemoattraction, apoptosis, etc. A message is received through a specific interaction of the cytokine with its corresponding receptor located on the plasma membrane of the receiving cell. This interaction triggers effector mechanisms within the responding cell which normally results in a beneficial outcome.


Disturbances in this communication pathway often cause ailments of varying severity. For example, a failure of immune cells to either produce or respond to interferons, a family of cytokines that indices antiviral and antiproliferative activities, may lead to diseases ranging from a mild viral infection to a lethal malignancy. In other cases, cellular communication imbalances or defective cytokine receptors may prevent normal control of cell growth, resulting in an overgrowth of a particular cell type that, thus, becomes cancerous to the body.


The discovery of new cytokines and the elucidation of their role in health and disease has become an area of extensive investigation in which remarkable advances have been made during the past five years. The availability of recombinant cytokines in significant quantities has stimulated both basic life science research and clinical studies aimed toward the development of improved tools to combat human diseases.

Before releasing our products for sale they are thoroughly analyzed to ensure the following qualities:
(NOTE:See indivdiual spec sheet with each shipment forbact specific information).

1. Authenticity: by N-terminal sequence analysis, amino acid composition, and, when possible, by SDS-PAGE, RP-HPLC, and FPLC analyses versus standards.


2. Purity: by SDS-PAGE, RP-HPLC, and FPLC analyses.


3. Biological Activity: by the relevant in vitro or in vivo bioassay.

4. Protein content: by UV spectroscopy, SDS-PAGE analysis, Lowry assay, and, when possible, by HPLC analysis versus the calibrated standard solution.


5. Endotoxin contamination: by kinetic LAL method.


6. Microbiological contamination: by membrane filtration method; protein solutions are sterile-filtered prior to vialing.

The relevant information relating to each product appears on the data sheet that is shipped with product. Please read this information carefully to obtain useful instructions for reconstitution and storage. If, after reading the data sheet, you need additional information, please review the following set of questions and answers, or call our technical service department.

1. WHAT SHOULD I KNOW ABOUT THE STABILITY OF YOUR PROTEIN PRODUCTS?

Unless otherwise mentioned on the product information sheet, all of our products are formulated in such a manner that the lyophilized proteins are very stable at room temperature. However, we recommend that for short-term storage (up to one month) they are best stored at 4'C. For longer periods, we recommend storing the lyophilized products at -20'C. We do not recommend storage at temperature below -50'C.

For reconstituted solutions of most products, we recommend short-term storage at 4'C. For longer term storage the protein solution should first be aliquoted (to avoid more than one freeze/thaw cycle) and stored frozen at -20'C. Please keep in mind that every freeze/thaw cycle may cause some denaturation of the protein.


2. WHY CAN'T I SEE THE PROTEIN PELLET IN THE VIAL?

Unlike many protein products available on the market, most RDI products are not formulated with carrier protein or other additives (e.g. BSA, HSA, sucrose,etc.) and are often lyophilized with a minimum amount of salt. As a result, the small amounts of protein can be deposited on the vial during lyophilization as a thin and, sometimes, invisible film. Before opening, we recommend centrifuging each vial in a microcentrifuge for 20-30 seconds to drive any protein that may be lodged in the cap or on the side to the bottom of the vial. Our quality control procedures assure that each vial contains the correct amount of product.

3. DO YOUR PRODUCTS CONTAIN ANY CARRIER PROTEINS OR OTHER ADDITIVES?

RDI does not add carrier protein or other bulk additives because we understand that many customers prefer preparation without them. Most of our products are also nearly salt-free, although some require the presence of particular buffers or salt concentrations for stability. This information is included in the formulation section of the product data sheet.


4. WHAT IS THE RELATIONSHIP BETWEEN THE SPECIFIC ACTIVITY EXPRESSED AS AN ED(50) AND AS UNITS/MG?

The ED(50) is defined as the cytokine concentration at which the activity is 50% of the maximum response. This method of expressing potency should only be used for cytokines whose dose-response curves are sigmoidal in shape. The formula for converting the activity as an ED(50) in ng/ml to specific activity in units/mg is:

       1 X 10(6)                = specific activity (units/mg)

      ED(50) (ng/ml)



5. WHICH CYTOKINES SHOW CROSS-SPECIES ACTIVITY?

With a few exceptions, most human cytokines are active on mouse cells. Many mouse cytokines are active on human cells, by may show lower specific activity than the corresponding human cytokine. A few human cytokines, such as IL-7, even exhibit higher specific activities on mouse cells than do the corresponding mouse cytokines. The interferons, GM-CSF,IL-3, and IL-4 are known to be species-specific with very little, if any, activity on non-homologous cells.

In contrast, the FGF's and neurotrophins are very highly conserved and show excellent activity on cells of other animal species.


6. HOW SHOULD I RECONSTITUTE THE VIAL OF LYOPHILIZED CYTOKINE?

Unless otherwise stated on the data sheet, RDI cytokines are formulated to be reconstituted in sterile water at 0.1-1 mg/ml. Most of our products are lyophilized from protein solutions without buffers and salts. However, a few cytokines have specific salt or pH requirements and these products are lyophilized from an appropriate formulation so that addition of water to the lyophilized protein restores these conditions. Additionally, the solubility of a few products is improved if they are dissolved in solutions of a specific composition. It is extremely important to follow the reconstitution instructions described on the product data sheet to ensure proper formulation of the cytokine. Once a cytokine solution has been prepared according to the recommendations, the user can make further dilutions into buffers appropriate to the particular biological application. Certain additives in solutions used for reconstitution may adversely affect the solubility and bioactivity of cytokines and should be tested in small scale before use.

-copyright 1997 by owner

-see our line of recombinant cytokines and corresponding antibodies

-see our line of human cytokine kits

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RDI Division of Fitzgerald Industries Intl

34 Junction Square Drive

Concord MA 01742-3049

USA

phone (800) 370-2222

      or (201) 584-7093   after June 1, 1997: (978) 371-6446 or (800) 370-2222

fax     (978) 371-2266    after June 1, 1997: (978) 371-2266

EMAIL:antibodies@fitzgerald-fii.com

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