Systems, compositions, and methods for transplantation and treating conditions

Abstract

Systems and methods for purification and concentration of autologous alpha-2 macroglobulin (A2M) from whole blood and or recombinant A2M are provided. Also provided are methods of treating wounds with A2M. Methods for utilizing A2M in combination with other treatments (e.g., platelets and other growth factors) are provided in addition to combinations with exogenous drugs or carriers. Also provided is a method of producing recombinant A2M wild type or variants thereof where the bait region was modified to enhance the inhibition characteristics of A2M and/or to prolong the half-life of the protein for treating wounds.

1 .- 390 . (canceled) 
     
     
         391 . A system for manual flow of a biological sample comprising a first module disposed above and in fluid connection with a second module:
 (a) the first module comprising:
 (i) a first retentate chamber with a first inlet, a second inlet, and a flow path therebetween; 
 (ii) a first permeate chamber with a first outlet; 
 (iii) a membrane between the first retentate chamber and the first permeate chamber; 
 (iv) a first and a second injector, 
   (b) the second module comprising:
 (i) a second retentate chamber with a third inlet, a fourth inlet, and a flow path therebetween; 
 (ii) a second permeate chamber; 
 (iii) a membrane between the second retentate chamber and the second permeate chamber; 
 (iv) a third and a fourth injector, 
   
       wherein the membrane of the first module has a pore size of from 0.1-2.0 microns, and the membrane of the second module has a pore size of from 10-500 kDa, and wherein the first, second, third, and fourth injector each comprises a manual actuator in fluid connection with the flow paths therebetween. 
     
     
         392 . The system of  claim 391 , wherein the first permeate chamber is in fluid communication with the second retentate chamber. 
     
     
         393 . The system of  claim 391 , wherein the first injector is connected to the first inlet via an adapter, and the second injector is connected to second inlet via an adapter, wherein the adapters are each independently a valve. 
     
     
         394 . The system of  claim 393 , wherein the first module further comprises a first vessel connected to the first inlet via the adaptor. 
     
     
         395 . The system of  claim 394 , wherein the first vessel comprises a syringe. 
     
     
         396 . The system of  claim 395 , wherein the first vessel is disposed below the first membrane, disposed above the first membrane, or oriented perpendicular to a surface of the membrane of the first module. 
     
     
         397 . The system of  claim 391 , further comprising a first and a second permeate collection reservoir. 
     
     
         398 . The system of  claim 397 , wherein the first permeate collection reservoir stores permeate from the first permeate chamber and retantate from the second retantate chamber, and wherein the second permeate collection reservoir stores permeate from the second permeate chamber. 
     
     
         399 . The system of  claim 391 , wherein the third injector is connected to the third inlet via an adapter, and the fourth injector is connected to the fourth inlet via an adapter, wherein the adapters are each independently a valve. 
     
     
         400 . The system of  claim 399 , further comprising a second vessel connected to the third inlet via the adapter. 
     
     
         401 . The system of  claim 400 , wherein the second vessel comprises a syringe. 
     
     
         402 . The system of  claim 401 , wherein the second vessel is disposed below the first membrane, disposed above the first membrane, or oriented perpendicular to a surface of the membrane of the second module. 
     
     
         403 . The system of  claim 391 , wherein the second permeate chamber comprises a second outlet, and wherein the second outlet is in fluid communication with a container or a bag via a second outlet flow path. 
     
     
         404 . The system of  claim 403 , wherein the second outlet flow path is in fluid connection with a connecting path, via a valve. 
     
     
         405 . The system of  claim 404 , wherein the second outlet flow path and the connecting flow path form a fluid communication between the second permeate chamber and the first retentate chamber. 
     
     
         406 . The system of  claim 405 , wherein the valve allows fluid to flow from the second outlet flow path to the container or bag in a first orientation, and wherein the valve allows fluid to flow from the container or bag to the first retentate chamber in a second orientation. 
     
     
         407 . The system of  claim 391 , wherein the second permeate chamber comprises a second outlet, and wherein the second outlet is in fluid connection with a collection vessel adapted to fit an adapter connected to the first or second inlet. 
     
     
         408 . The system of  claim 407 , wherein the collection vessel is the first or second injector. 
     
     
         409 . A method for enrichment of A2M from a biological sample obtained from an animal comprising:
 (a) flowing a biological sample through a system comprising: a first module disposed above and in fluid connection with a second module:
 (i) the first module comprising:
 (A) a first retentate chamber with a first inlet, a second inlet, and a flow path therebetween; 
 (B) a first permeate chamber with a first outlet; 
 (C) a membrane between the first retentate chamber and the first permeate chamber; 
 (D) a first and a second injector, 
 
 (ii) the second module comprising:
 (A) a second retentate chamber with a third inlet, a fourth inlet, and a flow path therebetween; 
 (B) a second permeate chamber with a second outlet; 
 (C) a membrane between the second retentate chamber and the second permeate chamber; 
 (D) a third and a fourth injector, 
 
   
       wherein the membrane of the first module has a pore size of from 0.1-2.0 microns, and the membrane of the second module has a pore size of from 10-500 kDa, and wherein the first, second, third, and fourth injector each comprises a manual actuator in fluid connection with the flow paths therebetween, and
 (b) collecting the retentate in the third or the fourth injector, wherein the retentate is enriched for A2M. 
 
     
     
         410 . The method of  claim 409 , wherein the biological sample is flown through the system by activation of one or more of the first, second, third, and fourth injectors.