Method for production of stroma-free hemoglobin

A technology of hemoglobin and blood, applied in the system field, can solve problems such as pollution and high cost, and achieve the effect of low-cost collection

Inactive Publication Date: 2002-01-23
桑伽特股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Second, the product must be safe
(See Winslow and Chapman, "Pilot Batch Preparation of Hemoglobin Solutions", Methods in Enzymology, 231:3-16, 1994, the disclosure of which is hereby incorporated by reference.) In

Method used

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  • Method for production of stroma-free hemoglobin
  • Method for production of stroma-free hemoglobin
  • Method for production of stroma-free hemoglobin

Examples

Experimental program
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Effect test

Embodiment 1

[0028] An experimental apparatus was constructed for the production of stroma-free hemoglobin from blood, which included a collection of emptied bags for expired blood; a cross-filter unit for washing, lysing, and purification; a bioreactor for cross-linking hemoglobin; preparative-grade high-efficiency liquid a phase chromatography (HPLC) instrument; and a fume hood where the product is finally filled into sterile bags. The flow circuits, including the PFW dispersion circuit, are sealed. All fittings are sanitary three-clamp (through) type, manufactured in a laminar flow environment. Excluding the office, the complete process takes about 1000ft 2 laboratory space. The capacity of the pilot plant was about 5 liters per week and the product cost was about $1000 / liter.

[0029] The cross filtration system consists of three pumps, four tanks and 4 different filter components. All construction materials are constructed of 316L stainless steel with an internal finish of 180 gri...

Embodiment 2

[0040] In an embodiment of the present invention, a self-contained portable device designed to collect donor / patient blood at the bedside is used. Suitable systems for this use include the Haemonetics MCS+8150 Blood Collection System (Haemonetics Corporation, Braintree, MA) and the COBE 2991 Cell Processor (COBE Laboratories, Inc., Lakewood, CO). Blood was collected in a CP2D / AS-3 anticoagulant / supplement system with a ratio of anticoagulant to anticoagulated blood of 1:16. The machine is configured to collect one or two units of red blood cells from a donor in about 25 minutes. Each unit collected yielded approximately 180 ml packed red blood cells ("RBC") and 400 ml plasma. The efficiency of the separation can be determined using the following relationship:

[0041] RBC count after treatment × weight after treatment × 100 = % RBC recovery

[0042] RBC count before treatment × weight before treatment

[0043] The number of lymphocytes remaining in the packed red blood cel...

Embodiment 3

[0067] Example 3: Preparation of modified hemoglobin using the described invention

[0068] The present invention can also be used in Figure 4 A modified hemoglobin solution ("product") is prepared from the stroma-free hemoglobin (SFH) solution prepared in the modified cell separator device shown.

[0069] Red blood cells can be obtained from any source, including animal or human, stored or fresh, or even expired human (blood) units obtained from blood banks. The improved procedure can also be used with hemoglobin solutions (including recombinant hemoglobin) prepared by any method, including the present invention or other methods. If donor blood is used, it is first mixed with an anticoagulant 50 , washed with saline 54 in a centrifuge rotor 52 , and lysed with distilled water 56 . The plasma, platelet and white blood cell fractions are removed and stored in separate compartments 58. A solution of hemoglobin (SFH) 60 is placed in a separate compartment reservoir 62 or cent...

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Abstract

The method employs a commercially-available blood cell separator comprising a computer-controlled centrifuge (2) having a rotor into which a blood processing bag (6) containing donor blood is placed. Once the blood is collected, the process is performed entirely within the enclosed centrifuge bowl, preferably in situation at the donor collection site. In the first step, the blood is centrifuged to separate the plasma from the cellular components. After isolation of the red blood cells from other blood components, the red cells are washed with normal saline or other solution. The red blood cells are then lysed by hypotonic shock to separate the red cell membranes (stroma) and the lysate is collected in a sterile container (44), leaving only the stroma in the centrifuge bowl. The final product can be used as raw material for any of the hemoglobin-based oxygen carriers currently being developed as red cell substitutes. All of the steps are performed within a processing container or blood bag (6) in the bowl centrifuge to minimize handling and maintain sterility. A method for preparing a modified hemoglobin solution incorporates the steps for producing stroma-free hemoglobin, then adding pre-measured reagents to react with the solution and filtering the solution.

Description

[0001] related application [0002] This application claims priority to Provisional Application No. 60 / 104,319, filed October 15, 1998, and Serial No. 60 / 122,180, filed March 1, 1999. The disclosure of the mentioned provisional application is hereby incorporated by reference. field of invention [0003] The present invention relates to systems and methods for the use of automated blood cell separators to prepare high-quality hemoglobin solutions as raw materials for the manufacture of hemoglobin-based therapeutic oxygen carriers ("blood substitutes"). Background of the invention [0004] Transfusion of stored human blood is an ancient backup method of medical practice. However its efficacy has never been shown strictly speaking, and the method has obvious drawbacks. For example, even in the best medical centers, when a blood transfusion is determined to be needed, treatment is delayed by the need...

Claims

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Application Information

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IPC IPC(8): A61L2/18A61K35/14A61M1/02A61M1/36A61P7/08B04B5/00
CPCA61M2001/3692A61M2001/3696A61M2202/0433A61M2205/331A61M2001/3698A61M2205/3317A61M1/3693A61M1/3692A61M1/3696A61M1/3698A61P7/08A61M2202/0071C07K14/805
Inventor R·M·温斯洛K·D·范德格里夫
Owner 桑伽特股份有限公司
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