1. 1&3: hGH-sc(Fc)2 2&4: hGH-Fc anti-Fc anti-hGH 1 2 3 4
Supplementary Data Figure 1. Identification of hGH fusion proteins. Both anti-hIgG (Fc specific) antibody and anti-hGH antibody recognized the target bands on the Western blot membrane of purified samples of hGH-sc(Fc)2 and hGH-Fc.

2. Supplementary Data Figure 2
Supplementary Data Figure 2. The hGH-Fc and hGH-sc(Fc)2 fusion proteins were analyzed for degradation following storage in PBS at 4 ˚C by SDS-PAGE/Coomassie blue staining. Sample identities are listed in the table blow.
Supplementary Data Table 1. Stability of hGH-sc(Fc)2 and hGH-Fc in PBS at 4 ˚C
Sample Lane #
Gel A
Gel B
Gel C
Sample
Days of storage 1
hGH-Fc 7
hGH-sc(Fc)2 6 2 17 16 3 28 27 4 35 34 5 47 48 59 60 73 74 8 88 89

3. Mannitol phosphate buffer
Supplementary Data Table 2. Stability of hGH-sc(Fc)2
Storage Conditions
Residual Activity*
Duration
Temp.
Buffer
0 month --
PBS
1 month
4 ˚C
106%
2 months
94%
-80 ˚C
74%
Mannitol phosphate buffer
*Activity was determined by the Nb2 cell proliferation assay and the ED50 values were compared.

4. Supplementary Data Figure 3. Characterization of hIgG1-Fc
Supplementary Data Figure 3. Characterization of hIgG1-Fc. Conditioned media containing hIgG1-Fc from transiently transfected HEK293 cells was collected and (A) analyzed by non‐reducing SDS‐PAGE and (B) reducing SDS-PAGE followed by Western blot probed with goat anti‐hIgG Fc specific antibody (1:3000 dilution), or (C) purified by Protein A‐Sepharose® 4B followed by native gel with Coomassie blue staining.
55-
70-
25-
35- A B C

5. 1 2 1: hGH-sc(Fc)2 2: sc(Fc)2-hGH 95 kDa- 72 kDa- 55 kDa- 43 kDa-
Supplementary Data Figure 4. Expression test of hGH fusion proteins. Anti-hGH antibody recognized the target bands on the Western blot membrane of the expression samples of hGH-sc(Fc)2 and sc(Fc)2-hGH. The expression level of hGH-sc(Fc)2 was about 3-fold higher than that of sc(Fc)2-hGH.
1: hGH-sc(Fc)2
2: sc(Fc)2-hGH
95 kDa-
72 kDa-
55 kDa-
43 kDa-
34 kDa-
26 kDa-