<p>INDEX CYTOKINE (Cat #) PAGE # HUMAN CYTOKINES 1 Endothelial cell growth supplement (ECGS) BD Biosciences Cat # 1 356006 Recombinant human /mouse/rat Activin A (R&D 338-AC) 1 Recombinant human Activin B (R&D Cat # 659-AB) 1 Recombinant human BMP4 (R&D 314BP) 2 Recombinant human BMPR-1A/ALK-3/Fc Chimera (R&D Cat # 2 315-BR) Recombinant human BMPR-1B/ALK-6/Fc Chimera (R&D Cat # 2 315-BR) Recombinant human Dkk1 (R&D Cat # 1096-DK) 3 Recombinant human EGF (R&D Cat# 236-EG) 3 Recombinant human EPO (Amgen–Sinai Pharmacy) 3 Recombinant human FGF2 (R&D 233-FB) 4 Recombinant human FGF4 (R&D Cat # 235-F4) 4 Recombinant human GM-CSF (R&D Cat # 215-GM) 5 Recombinant human HGF (R&D Cat # 294-HG) 5 Recombinant human IGF-1 (R&D 291-G1) 5 Recombinant human IGF-II (R&D Cat # 292-G2) 6 Recombinant human IL-3 (R&D Cat # 203-IL) 6 Recombinant human IL-6 (R&D 206-IL) 6 Recombinant human IL-11 (R&D Cat # 218-IL) 7 Recombinant human M-CSF (R&D Cat # 216-MC) 7 Recombinant human Netrin (R&D Cat # 1254-N4) 7 Recombinant human Osteopontin (R&D Cat # 1433-OP) 8 Recombinant human SCF (R&D 255-SC) 8 Recombinant human Sonic Hedgehog (R&D Cat # 1845-SH) 8 Recombinant human TPO (R&D Cat # 288-TP) 9 Recombinant human VEGF (R&D Cat # 293-VE) 9 MOUSE CYTOKINES Recombinant mouse DLL4 (R&D Cat # 1389-D4) 10 Recombinant mouse EGF (R&D Cat# 2028-EG) 10 Recombinant mouse GM-CSF (R&D Cat # 415-ML) 10 Recombinant mouse IGF-I (R&D Cat # 791-MG) 11 Recombinant mouse IL-11 (R&D Cat # 418-ML) 11 Recombinant mouse LIF (R&D Cat# 449-L) 11 Recombinant mouse M-CSF (R&D Cat # 416-ML) 12 Recombinant mouse Sonic Hedgehog (R&D Cat # 461-SH) 12 Recombinant mouse TPO (R&D Cat # 488-TO) 12 Recombinant mouse Wnt 3A (R&D Cat # 1324-WN/CF) 13 Recombinant mouse Wnt 5A (R&D Cat # 645-WN) 13 RECONSTITUTING CYTOKINES</p><p>1 Endothelial cell growth supplement (ECGS) BD Biosciences Cat # 356006 – 100mg/vial.</p><p> In a 14 ml tube, add 10 ml of IMDM reconstituted with glutamine and P/S.</p><p> Using 5 ml pipette, transfer ` 2-3 ml of this media into the vial containing lyophilized ECGS.</p><p> Let it dissolve at RT for 30 minutes. Mix gently. </p><p> Aliquot 2000uL / 5 ml tube (5 tubes). Label tubes with date and concentration of ECGS (10mg/ml ECGS). </p><p> Store aliquots at -20.</p><p>Recombinant human/mouse/rat Activin A (R&D Cat # 338-AC) – 250ug</p><p> In a 14 ml tube, add 5 ml of 0.1% BSA in 1X PBS previously filtered.</p><p> Using 10ml pipette, transfer some (`1ml) of this media into the vial containing lyophilized hActivin A</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 14ml tube containing the remaining 4 ml of media and mix gently with pipette.</p><p> Aliquot 500uL / eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of hActA (50ug/ml hActA). </p><p> Store at –20.</p><p>Recombinant human Activin B (R&D Cat # 659-AB) – 250ug (NOT IN STOCK)</p><p> In a 14 ml tube, add 5 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 10ml pipette, transfer some (`1ml) of this media into the vial containing lyophilized hActivinB</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 14ml tube containing the remaining 4 ml of media and mix gently with pipette.</p><p> Aliquot 500uL / eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of hActB (50ug/ml hActB). </p><p>2 Store at –20.</p><p>Recombinant human BMP4 (R&D Cat# 314-BP) – 250ug Reagents: 10% BSA, 1N HCL and Sterile water</p><p> In a 50 ml tube, add 30ml of sterile water, 300uL of 10% BSA and 120uL of 1N HCL</p><p> Filter this mixture</p><p> Transfer 25ml of the above media into another 50ml tube.</p><p> Using 10ml pipette, transfer some of this media (2-5ml) into the vial containing lyophilized hBMP4</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 50 ml tube containing the remaining 20-23 ml of media and mix gently with pipette.</p><p> Aliquot 500uL / eppendorf (50 eppendorfs). Label eppendorfs with date and concentration of hBMP4 (10ug/ml hBMP4). </p><p> Store at –20.</p><p>Recombinant human BMPR-1A/ALK-3/Fc Chimera (R&D Cat # 315-BR) – 100ug</p><p> Using a 1 ml pipette, transfer 1ml of 0.1% BSA in 1X PBS, previously filtered into the vial containing lyophilized hBMPR-1A.</p><p> Let it dissolve at RT for 30 minutes. Mix gently.</p><p> Aliquot 100uL / eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of hBMPR-1A (100ug / ml hBMPR-1A). Store at –20.</p><p>Recombinant human BMPR-1B/ALK-6/Fc Chimera (R&D Cat # 505-PR) – 100ug</p><p> Using a 1 ml pipette, transfer 1ml of 0.1% BSA in 1X PBS, previously filtered into the vial containing lyophilized hBMPR-1B.</p><p> Let it dissolve at RT for 30 minutes. Mix gently.</p><p>3 Aliquot 100uL / eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of hBMPR-1B (100ug /ml BMPR-1B). Store at –20.</p><p>Recombinant human Dkk1 (R&D Cat # 1096-DK) – 250ug</p><p> In a 5 ml tube, add 2.5 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 5 ml pipette, transfer media into the vial containing lyophilized hDKK1.</p><p> Let it dissolve at RT for 30 minutes. Mix gently. </p><p> Aliquot 200uL / eppendorf (13 eppendorfs). Label eppendorfs with date and concentration of hDKK1 (100ug /ml hDkk1). Store at –20.</p><p>Recombinant human EGF (R&D Cat# 236-EG) – 250ug Reagents: 1M acetic acid and 0.1% BSA in 1XPBS </p><p> In a 50 ml tube, add 30ml of 1X PBS containing 0.1% BSA.</p><p> To this media add 300ul (10uL/ml) of a 1M acetic acid solution. </p><p> Filter this mixture</p><p> Transfer 25ml of the above media into another 50ml tube.</p><p> Using 10ml pipette, transfer some of this media (2-5ml) into the vial containing lyophilized hEGF</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 50 ml tube containing the remaining 20-23 ml of media and mix gently with pipette.</p><p> Aliquot 1000uL / eppendorf (25 eppendorfs). Label eppendorfs with date and concentration of hEGF (10ug/ml hEGF). </p><p> Store at –20.</p><p>Recombinant human EPO (Amgen – Sinai Pharmacy) – 10,000U/ml</p><p> In a 14 ml tube, add 4 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Remove metal cap. Insert sterile needle attached to a sterile 1ml syringe into the rubber cap of the vial.</p><p>4 Aspirate out the 1ml of hEPO and add it to the 4ml of 0.1% BSA in 1X PBS. Mix gently.</p><p> Aliquot 1000uL / eppendorf (5 eppendorfs). Label eppendorfs with date and concentration of hEPO (2000U/ml hEPO). </p><p> Store at 4 degrees. Do not freeze.</p><p>Recombinant human bFGF2 (R&D Cat # 233-FB) – 250ug</p><p> In a 50 ml tube, add 30ml of 0.1% BSA in 1X PBS </p><p> To the above media, add 300uL of 0.1M DTT </p><p> Filter this mixture</p><p> Transfer 25ml of the above media into another 50ml tube.</p><p> Using 10ml pipette, transfer some of this media (2-5ml) into the vial containing lyophilized human bFGF</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 50 ml tube containing the remaining 20-23 ml of media and mix gently with pipette.</p><p> Aliquot 1000uL / eppendorf (25 eppendorfs). Label eppendorfs with date and concentration of human bFGF (10ug/ml bFGF). </p><p> Store at –20.</p><p>Recombinant human FGF4 (R&D Cat # 235-F4) – 25ug</p><p> In a 5 ml tube, add 1 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Transfer the 1 ml of the above media into another the lyophilized FGF4 vial.</p><p> Let it dissolve at RT for 30 minutes. Mix gently with pipette.</p><p> Aliquot 200uL / eppendorf (5 eppendorfs). Label eppendorfs with date and concentration of bFGF4 (10ug/ml FGF4). </p><p>Recombinant human GM-CSF (R&D Cat # 215-GM) – 10ug </p><p>5 In a 14 ml tube, add 10 ml of 0.1% BSA in 1X PBS </p><p> Using 10ml pipette, transfer some of this media (1-2 ml) into the vial containing lyophilized hGM-CSF</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 14 ml tube containing the remaining 9-8 ml of media and mix gently with pipette.</p><p> Aliquot 500uL / eppendorf (20 eppendorfs). Label eppendorfs with date and concentration of hGM-CSF (1 ug / ml bFGF). </p><p>Recombinant human HGF (R&D Cat # 294-HG) – 250ug</p><p> In a 5 ml tube, add 2.5 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 5 ml pipette, transfer all this media into the vial containing lyophilized hHGF.</p><p> Let it dissolve at RT for 30 minutes. Mix gently. </p><p> Aliquot 200uL / eppendorf (13 eppendorfs). Label eppendorfs with date and concentration of hHGF (100ug / ml hHGF). Store at –20.</p><p>Recombinant human IGF-I (R&D Cat # 291-G1) – 250ug</p><p> In a 14 ml tube, add 10 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 10ml pipette, transfer some (1-2ml) of this media into the vial containing lyophilized hIGF-I.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 14 ml tube containing the remaining 8-10 ml of media and mix gently with pipette.</p><p> Aliquot 500uL / eppendorf (20 eppendorfs). Label eppendorfs with date and concentration of hIGF-I (25ug/ml hIGF-I). </p><p> Store at –20.</p><p>Recombinant human IGF-II (R&D Cat # 292-G2) – 25 ug</p><p>6 In a 5 ml tube, add 2.5 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 1 ml pipette, transfer some (1-2ml) of this media into the vial containing lyophilized hIGF-II.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 5 ml tube containing the remaining 0.5-1.5 ml of media and mix gently with pipette.</p><p> Aliquot 250uL / eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of hIGF-II (10 ug/ml hIGF-II). </p><p> Store at –20.</p><p>Recombinant human IL-3 (R&D Cat # 203-IL) – 50 ug </p><p> In a 14 ml tube, add 5 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 1 ml pipette, transfer some (1-2ml) of this media into the vial containing lyophilized hIL-3.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 14 ml tube containing the remaining 3-4 ml of media and mix gently with pipette.</p><p> Aliquot 500 uL / eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of hIL-3 (10 ug /ml, hIL-3). </p><p> Store at –20.</p><p>Recombinant human IL-6 (R&D Cat # 206-IL) – 50ug</p><p> In a 14 ml tube, add 5 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 10ml pipette, transfer 1ml of this media into the vial containing lyophilized hIL-6</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 14ml tube containing the remaining 4 ml of media and mix gently with pipette.</p><p> Aliquot 500uL / eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of hIL-6 (10ug/ml, hIL-6). Store at –20.</p><p>Recombinant human IL-11 (R&D Cat # 218-IL) – 250ug</p><p>7 In a 14 ml tube, add 5 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 10ml pipette, transfer 1ml of this media into the vial containing lyophilized hIL-11.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 14ml tube containing the remaining 4 ml of media and mix gently with pipette.</p><p> Aliquot 500uL / eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of hIL-11 (50ug/ml, hIL-11). Store at –20.</p><p>Recombinant human M-CSF (R&D Cat # 216-MC) – 25ug </p><p> In a 5 ml tube, add 2.5 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 1 ml pipette, transfer some of this media (1-2 ml) into the vial containing lyophilized hM-CSF.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 5 ml tube containing the remaining 0.5-1.5 ml of media and mix gently with pipette.</p><p> Aliquot 500uL / eppendorf (5 eppendorfs). Label eppendorfs with date and concentration of hM-CSF (10 ug /ml hM-CSF). </p><p>Recombinant human Netrin (R&D Cat # 1254-N4) – 25 ug </p><p> In a 5 ml tube, add 2.5 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 1 ml pipette, transfer 1 ml of this media into the vial containing lyophilized hNetrin.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 5 ml tube containing the remaining 1.5 ml of media and mix gently with pipette.</p><p> Aliquot 250 uL / eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of hNetrin (10 ug /ml hNetrin). </p><p> Store at –20.</p><p>Recombinant human Osteopontin (R&D Cat # 1433-OP) – 50 ug </p><p>8 In a 14 ml tube, add 5 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 1 ml pipette, transfer 1 ml of this media into the vial containing lyophilized hOsteopontin.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 14 ml tube containing the remaining 4 ml of media and mix gently with pipette.</p><p> Aliquot 500 uL / eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of hOsteopontin (10 ug /ml hOsteopontin). </p><p> Store at –20.</p><p>Recombinant human SCF (R&D Cat # 255-SC) – 50ug</p><p> In a 5 ml tube, add 1 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 1 ml p1000, transfer 1ml of this media into the vial containing lyophilized hSCF.</p><p> Let it dissolve at RT for 30 minutes. Mix gently.</p><p> Aliquot 500uL / eppendorf (2 eppendorfs). Label eppendorfs with date and concentration of hSCF (50ug/ml hSCF). </p><p> Store at –20.</p><p>Recombinant human Sonic Hedgehog (R&D Cat # 1845-SH) – 50 ug </p><p> Using p1000, transfer 250ul of 1X PBS containing 0.1% BSA into the vial containing lyophilized hShh.</p><p> Let it dissolve at RT for 30 minutes and mix gently with pipette.</p><p> Aliquot 50 uL / eppendorf (5 eppendorfs). Label eppendorfs with date and concentration of hShh (200 ug/ml hShh). </p><p> Store at –20.</p><p>Recombinant human TPO (R&D Cat # 288-TP) – 250 ug </p><p>9 In a 50 ml tube, add 25ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 10ml pipette, transfer some of this media (2-5ml) into the vial containing lyophilized TPO</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 50 ml tube containing the remaining 20-23 ml of media and mix gently with pipette.</p><p> Aliquot 1000uL / eppendorf (25 eppendorfs). Label eppendorfs with date and concentration of hTPO (10ug/ml hTPO). </p><p>Recombinant human VEGF 165 (R&D Cat # 293-VE) – 50 ug </p><p> In a 14 ml tube, add 10 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 10ml pipette, transfer some (1-2ml) of this media into the vial containing lyophilized hVEGF.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 14 ml tube containing the remaining 8-10 ml of media and mix gently with pipette.</p><p> Aliquot 500uL / eppendorf (20 eppendorfs). Label eppendorfs with date and concentration of hVEGF (5ug/ml hVEGF). Store at –20.</p><p>10 MOUSE CYTOKINES</p><p>Recombinant mouse DLL4 (R&D Cat # 1389-D4) – 50 ug </p><p> In a 5 ml tube, add 2.5 ml of 0.1% BSA in 1X PBS, previously filtered. </p><p> Using a p1000, transfer 1 ml of this media into the vial containing lyophilized mDLL4.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 5 ml tube containing the remaining 1.5 ml of media and mix gently with pipette.</p><p> Aliquot 500uL / eppendorf (5 eppendorfs). Label eppendorfs with date and concentration of mDLL4 (20ug/ml mDLL4). </p><p>Recombinant mouse EGF (R&D Cat# 2028-EG) – 200ug</p><p> Add 2 ml of 1X PBS containing 0.1% BSA to the vial containing lyophilized mEGF.</p><p> Let it dissolve at RT for 30 minutes. Mix gently. </p><p> Aliquot 500 uL / eppendorf (4 eppendorfs). Label eppendorfs with date and concentration of mEGF (100ug/ml mEGF). </p><p> Store at –20.</p><p>Recombinant mouse GM-CSF (R&D Cat # 415-ML) – 25ug </p><p> In a 5 ml tube, add 2.5 ml of 0.1% BSA in 1X PBS, previously filtered. </p><p> Using a p1000, transfer 1 ml of this media into the vial containing lyophilized mGM-CSF.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 5 ml tube containing the remaining 1.5 ml of media and mix gently with pipette.</p><p> Aliquot 500uL / eppendorf (5 eppendorfs). Label eppendorfs with date and concentration of mGM-CSF (10 ug/ml mGM-CSF). </p><p>11 Recombinant mouse IGF-I (R&D Cat # 791-MG) – 50 ug</p><p> In a 5 ml tube, add 2 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using a p1000 pipette, transfer 1ml of this media into the vial containing lyophilized mIGF-I.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 5 ml tube containing the remaining 1 ml of media and mix gently with pipette.</p><p> Aliquot 250uL / eppendorf (8 eppendorfs). Label eppendorfs with date and concentration of mIGF-I (25ug/ml mIGF-I). Use at 1uL/ml.</p><p> Store at –20.</p><p>Recombinant mouse IL-11 (R&D Cat # 418-ML) – 25 ug</p><p> In a 14 ml tube, add 5 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using a p1000 pipette, transfer 1ml of this media into the vial containing lyophilized mIL-11.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 14 ml tube containing the remaining 4 ml of media and mix gently with pipette.</p><p> Aliquot 500uL / eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of mIL-11 (5 ug/ml mIL-11). Use at 1uL/ml.</p><p> Store at –20.</p><p>Recombinant mouse LIF (R&D Cat# 449-L) – 50ug</p><p> In a 14 ml tube, add 5 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 1 ml pipette, transfer 1 ml into the vial containing lyophilized mLIF.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 14 ml tube containing the remaining 4 ml of media and mix gently with pipette.</p><p> Aliquot 500uL / eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of mLIF (10ug/ml mLIF). Use @ 1uL/ml for mESC maintenance in serum free media. Store at –20.</p><p>12 Recombinant mouse M-CSF (R&D Cat # 416-ML) – 50ug </p><p> In a 14 ml tube, add 5 ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 1 ml pipette, transfer 1 ml) into the vial containing lyophilized mM-CSF.</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 14 ml tube containing the remaining 4 ml of media and mix gently with pipette.</p><p> Aliquot 500uL /eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of mM-CSF (10ug/ml mM-CSF). </p><p>Recombinant mouse Sonic Hedgehog (R&D Cat # 461-SH) – 25 ugs </p><p> Using p1000, transfer 500ul of 1X PBS containing 0.1% BSA into the vial containing lyophilized mShh.</p><p> Let it dissolve at RT for 30 minutes and mix gently with pipette.</p><p> Aliquot 100 uL /eppendorf (5 eppendorfs). Label eppendorfs with date and concentration of mShh (50 ug/ml mShh). </p><p> Store at –20.</p><p>Recombinant mouse TPO (R&D Cat # 488-TO) – 50 ug </p><p> In a 50 ml tube, add 5ml of 0.1% BSA in 1X PBS, previously filtered.</p><p> Using 1 ml pipette, transfer 1 ml into the vial containing lyophilized mTPO</p><p> Let it dissolve at RT for 30 minutes. Transfer media back into 5 ml tube containing the remaining 4 ml of media and mix gently with pipette.</p><p> Aliquot 500uL /eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of mTPO (10 ug /ml mTPO). </p><p>13 Recombinant mouse Wnt 3A (R&D Cat # 1324-WN/CF)</p><p> Supplied as a 0.2uM filtered 5ml solution in 1X PBS containing 0.1mM EDTA, 0.5% CHAPS, pH 6.8 containing 200ug of Wnt3A @ a concentration of 40ng/ml. </p><p> Using p1000 pipette, aliquot 200uL / eppendorf (25 eppendorfs). Label eppendorfs with date and concentration of mWnt3A (40 ng/ml mWnt3A). </p><p>Recombinant mouse Wnt 5A (R&D Cat # 645-WN) – 10 ug </p><p> Using p1000 pipette, transfer 1ml of 1X PBS with 0.1% BSA, previously filtered into the lyophilized vial containing mWnt5A.</p><p> Let it dissolve at RT for 30 minutes. </p><p> Aliquot 100uL / eppendorf (10 eppendorfs). Label eppendorfs with date and concentration of mWnt5A (10 ug / ml mWnt5A). </p><p>14</p>
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