OPTIMIZATION METHOD FOR CAPTURING PROTEINS BY MULTI-COLUMN CONTINUOUS CHROMATOGRAPHY (MCC)

Abstract

An optimization method for capturing proteins by multi-column continuous chromatography (MCC), including the following steps: step 1, under the conditions of a set loading protein concentration and an arbitrary load residence time, performing a single time of protein breakthrough experiment to obtain a protein breakthrough curve; step 2, under a set breakthrough percentage for a target protein, integrating the breakthrough curve to obtain a single-column loading capacity and establishing a linear relationship between the interconnected load time and the load residence time; step 3, solving for the optimal number of operating columns for capturing proteins by MCC based on step 2; step 4, solving for the optimal load residence time for capturing proteins by MCC based on step 2, step 3; and step 5, solving for the maximum productivity of capturing proteins by MCC based on step 4.

Claims

1. An optimization method for capturing proteins by a multi-column continuous chromatography (MCC), wherein the MCC is used for a protein capture, and a number of columns is greater than or equal to 3, the optimization method comprises the following steps: step 1, under conditions of a set loading protein concentration and an arbitrary load residence time, performing a single time of a protein breakthrough experiment to obtain a protein breakthrough curve; step 2, under a set breakthrough percentage for a target protein, integrating the protein breakthrough curve to obtain a single-column loading capacity of the MCC as a first formula: $A=\frac{{\int }_{t=0}^{t_{l\_s}}\left[c_{\exp }-c\left(t\right)\right]\mathrm{dt}}{{\mathrm{RT}}_C}-c_{\exp }\times s$ in the first formula, s is the set breakthrough percentage for the target protein, A (g/L) is the single-column loading capacity of the MCC obtained by integrating the protein breakthrough curve at the set breakthrough percentage s, t is a loading time, t.sub.1_s (min) is a loading time until reaching the set breakthrough percentage s, and c.sub.exp (g/L) is a loading protein concentration; c(t) (g/L) is a breakthrough protein concentration, and RT.sub.C (min) is a single-column residence time of an interconnected load of the MCC; establishing a linear relationship between an interconnected load time t.sub.C and a load residence time RT.sub.C through the single-column loading capacity further comprises the following step: substituting the single-column loading capacity obtained in step 2 into a second formula $t_C=\frac{A\times {\mathrm{RT}}_C}{c_{\exp }}-t_{\mathrm{CW}},$ wherein t.sub.C (min) is the interconnected load time of the MCC, RT.sub.C (min) is the single-column residence time of the interconnected load of the MCC, A (g/L) is the single-column loading capacity of the MCC obtained in step 2 by integrating the protein breakthrough curve at the set breakthrough percentage for the targe protein, t.sub.CW (min) is an interconnected wash time of the MCC, and c.sub.exp (g/L) is the loading protein concentration; through the first formula and the second formula, the linear relationship between the interconnected load time t.sub.C and the load residence time RT.sub.C is obtained; step 3, solving an optimal number of operating columns for capturing the proteins by the MCC under the set loading protein concentration and a set protein breakthrough percentage based on the linear relationship between the interconnected load time t.sub.C and the load residence time RT.sub.C obtained in step 2 further comprises the following steps: drawing a line t.sub.C -RT.sub.C in a t-RT coordinate system based on the linear relationship between the interconnected load time t.sub.C and the load residence time RT.sub.C obtained in step 2, and drawing a line of a third formula $t=\frac{t_{\mathrm{RR}}-\left(N-3\right)t_{\mathrm{CW}}}{\left(N-2\right)}$ in the t-RT coordinate system, wherein t.sub.CW (min) is the interconnected wash time of the MCC, t.sub.RR (min) is a recovery and regeneration (R-R) time of the MCC and comprises a sum of a washing time, an elution time, and a regeneration time, and N is a number of the operating columns; by adjusting a N value, an intersection of two lines is changed so that a load residence time corresponding to the intersection is within a set residence time range; if only one N value meets above conditions, then the N value is the optimal number of the operating columns for capturing the proteins by the MCC under the set loading protein concentration and the set protein breakthrough percentage; if two or more N values meet the above conditions, a largest N value is selected as the optimal number of the operating columns; step 4, solving an optimal load residence time for the capturing proteins by the MCC under the set loading protein concentration, the set protein breakthrough percentage, and the optimal number of the operating columns based on the linear relationship between the interconnected load time t.sub.C and the load residence time RT.sub.C obtained in step 2; and step 5, solving a maximum productivity of capturing the proteins by the MCC based on the optimal load residence time obtained in step 4.

2. The optimization method for capturing the proteins by the MCC according to claim 1, wherein the set breakthrough percentage for the target protein is greater than or equal to 50%.

3. The optimization method for capturing the proteins by the MCC according to claim 1, wherein solving the optimal load residence time in step 4 comprises the following step: $t_C=\frac{A\times {\mathrm{RT}}_C}{c_{\exp }}-t_{\mathrm{CW}}$ solving simultaneous equations of the second formula of the linear relationship between the interconnected load time t.sub.C and the load residence time RT.sub.C obtained in step 2 and the third formula $t=\frac{t_{\mathrm{RR}}-\left(N-3\right)t_{\mathrm{CW}}}{\left(N-2\right)}$ to obtain the optimal load residence time, wherein t.sub.C (min) is the interconnected load time of the MCC, t.sub.CW (min) is the interconnected wash time of the MCC, t.sub.RR (min) is the R-R time of the MCC and comprises the sum of the washing time, the elution time, and the regeneration time, c.sub.exp (g/L) is the loading protein concentration, and N is the number of the operating columns; the optimal load residence time for capturing the proteins by the MCC under the set loading protein concentration, the set protein breakthrough percentage, and the optimal number of the operating columns is obtained by solving the simultaneous equations.

4. The optimization method for capturing the proteins by the MCC according to claim 1, wherein a graphic method used to solve the optimal load residence time in step 4 comprises the following steps: drawing the line t.sub.C -RT.sub.C in the t-RT coordinate system based on the linear relationship between the interconnected load time t.sub.C and the load residence time RT.sub.C obtained in step 2, and drawing the line of the third formula $t=\frac{t_{\mathrm{RR}}-\left(N-3\right)t_{\mathrm{CW}}}{\left(N-2\right)}$ in the t-RT coordinate system, wherein t.sub.CW (min) is the interconnected wash time of the MCC, t.sub.RR (min) is the R-R time of the MCC and comprises the sum of the washing time, the elution time, and the regeneration time, and N is the number of the operating columns; the load residence time corresponding to the intersection of the two lines is the optimal load residence time for capturing the proteins by the MCC under the set loading protein concentration, the set protein breakthrough percentage, and the optimal number of the operating columns.

5. The optimization method for capturing the proteins by the MCC according to claim 1, wherein solving the maximum productivity in step 5 comprises the following step: substituting the optimal load residence time obtained in step 4 into a fourth formula $P_{C,\mathrm{opt}}=\frac{c_{\exp }}{N\times {\mathrm{RT}}_{C,\mathrm{opt}}},$ wherein P.sub.C,opt (g/L/h) is the maximum productivity under the optimal load residence time, RT.sub.C,opt is the optimal load residence time obtained in step 4, c.sub.exp (g/L) is the loading protein concentration, and N is the number of the operating columns; the maximum productivity of capturing the proteins by the MCC is solved by the fourth formula.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0032] FIG. 1 is a schematic diagram of the operation mode of capturing proteins by a three-column continuous chromatography.

[0033] FIG. 2 is a schematic diagram of the operation mode of capturing proteins by a four-column continuous chromatography.

[0034] FIG. 3 shows a breakthrough curve obtained when the Praesto® Jetted A50 resin is used at the loading protein concentration of 5 g/L and the load residence time of 2 min.

[0035] FIG. 4 shows a solving method of the optimal number of operating columns when the Praesto® Jetted A50 resin is used at the loading protein concentration of 5 g/L.

[0036] FIG. 5 shows a breakthrough curve obtained when the Praesto® Jetted A50 resin is used at the loading protein concentration of 4 g/L and the load residence time of 4 min.

[0037] FIG. 6 shows a solving method of the optimal load residence time when the Praesto® Jetted A50 resin is used at the loading protein concentration of 4 g/L.

[0038] FIG. 7 shows a breakthrough curve obtained when the Mabselect SuRE™ LX resin is used at the loading protein concentration of 8 g/L and the load residence time of 3 min.

[0039] FIG. 8 shows a solving method of the optimal number of operating columns when the Mabselect SuRE™ LX resin is used at the loading protein concentration of 8 g/L.

[0040] FIG. 9 shows a breakthrough curve obtained when the Mabselect™ SuRE LX resin is used at the loading protein concentration of 6 g/L and the load residence time of 3 min.

[0041] FIG. 10 shows a solving method of the optimal load residence time when the Mabselect SuRE™ LX resin is used at the loading protein concentration of 6 g/L.

DETAILED DESCRIPTION OF THE EMBODIMENTS

[0042] The technical solutions of the present invention will be clearly and completely described below in combination with the embodiments and drawings of the present invention. Obviously, the embodiments described are part of the embodiments of the present invention, rather than all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those having ordinary skill in the art without creative labor are within the scope of protection of the present invention.

[0043] The present invention provides an optimization method for capturing proteins by MCC, where the MCC is used for protein capture, and the number of columns is greater than or equal to 3. The optimization method includes the following steps:

[0044] Step 1: Under the conditions of a set loading protein concentration and an arbitrary load residence time, a single protein breakthrough experiment is performed to obtain a protein breakthrough curve.

[0045] Step 2: Under a set breakthrough percentage (greater than or equal to 50%), the breakthrough curve is integrated to obtain a single-column loading capacity of MCC, and a linear relationship between the interconnected load time and the load residence time is established through the single-column loading capacity.

[0046] Step 3: The optimal number of operating columns for capturing proteins by MCC under the set loading protein concentration and the protein breakthrough percentage is solved based on the linear relationship between the interconnected load time and the load residence time obtained in step 2.

[0047] Step 4: The optimal load residence time for capturing proteins by MCC under the set loading protein concentration, the protein breakthrough percentage, and the optimal number of operating columns is solved based on the linear relationship between the interconnected load time and the load residence time obtained in step 2.

[0048] Step 5: The maximum productivity of capturing proteins by MCC is solved based on the optimal load residence time obtained in step 4.

[0049] After obtaining the above optimal parameters, the present invention adopts MPCC for protein capture according to the obtained optimal parameters. In a separation cycle, each column completes all steps of the continuous chromatography operation and returns to its initial state.

Embodiment 1. The Solution of the Optimal Number of Operating Columns

(1) The Breakthrough Curve Obtained by Experiments

[0050] The Praesto Jetted® A50 resin from Purolite company is used to pack a 5 ml chromatographic column. The immunoglobulin G with a concentration of 5 g/L is used for loading, and the load residence time is 2 min. The breakthrough experiment is conducted, and the loading is stopped when the breakthrough protein concentration reaches 4.5 g/L. The breakthrough curve is shown in FIG. 3.

(2) The Establishment of the Linear Relationship between the Interconnected Load Time and the Load Residence Time

[0051] When the set breakthrough percentage is 0.5 (that is, 50% breakthrough), the breakthrough curve is integrated to obtain the single-column loading capacity of 79.8 g/L. The R-R time t.sub.RR is 55 min and the interconnected wash time t.sub.CW is 8 min after optimization by conventional batch chromatography experiment. The above single-column loading capacity, etc., is substituted into the following formula, and the linear relationship between the interconnected load time t.sub.C and the load residence time RT.sub.C can be written as follows:

[00008]$t_C=\frac{A\times {\mathrm{RT}}_C}{c_{\exp }}-t_{\mathrm{CW}}=15.9{\mathrm{RT}}_C-8$

(3) The Solution of the Optimal Number of Operating Columns

[0052] According to the above linear relationship equation, the line t.sub.C -RT.sub.C is drawn in the t-RT coordinate system, as shown in FIG. 4.

[0053] According to the linear equation of

[00009]$t=\frac{t_{\mathrm{RR}}-\left(N-3\right)t_{\mathrm{CW}}}{\left(N-2\right)},$

when the number of columns N equals 3, then

[00010]$t=\frac{55-\left(3-3\right)\times 8}{\left(3-2\right)}=55\left(\min \right)$

[0054] In the t-RT coordinate system, line t=55 is drawn.

[0055] As shown in FIG. 4, the RT.sub.C of the intersection of t.sub.C -RT.sub.C and t=55 is 3.9 min. Generally, it is recommended that the single-column residence time in the interconnected load stage of MCC is 2 min-4 min, and this RT.sub.C is within this range. Therefore, this system is suitable for three-column continuous chromatography.

[0056] Let the number of columns N=4, there is:

[00011]$t=\frac{55-\left(4-3\right)\times 8}{\left(4-2\right)}=23.5\left(\min \right)$

[0057] In the t-RT coordinate system, line t=23.5 is drawn.

[0058] As shown in FIG. 4, the RT.sub.C of the intersection of t.sub.C -RT.sub.C and t=23.5 is 2 min, which is within the recommended load residence time range. Therefore, this system is suitable for the four-column MCC. Since the system applies to both the three-column system and the four-column system, the four-column system with more columns is selected to be the optimal operating system.

[0059] Under the conditions of the protein concentration being 5 g/L and the protein breakthrough percentage being 0.5, the three-column, four-column, and five-column MCC experiments are performed with the Praesto® Jetted A50 resin. The maximum productivity of the three modes is 25.6 g/L (the load residence time is 3.9 min), 37.5 g/L (the load residence time is 2 min), and 30 g/L (the load residence time is 2 min), respectively. Specifically, the four-column continuous chromatography has the maximum productivity, which is consistent with the optimization results, so the optimal number of operating columns is verified to be 4.

Embodiment 2. The Solution of the Optimal Load Residence Time

(1) The Breakthrough Curve Obtained by Experiments

[0060] The Praesto® Jetted A50 resin from Purolite company is used to pack a 5 ml chromatographic column. The immunoglobulin G with a concentration of 4 g/L is used for loading, and the load residence time is 4 min. The breakthrough experiment is conducted, and the loading is stopped when the breakthrough protein concentration reaches 3.6 g/L. The breakthrough curve is shown in FIG. 5.

(2) The Establishment of the Linear Relationship between the Interconnected Load Time and the Load Residence Time

[0061] When the set breakthrough percentage is 0.7 (that is, 70% breakthrough), the breakthrough curve is integrated to obtain the single-column loading capacity of 90.1 g/L. The R-R time t.sub.RR is 55 min and the interconnected wash time t.sub.CW is 8 min after optimization by conventional batch chromatography experiment. The above single-column loading capacity, etc., is substituted into the following formula, and the linear relationship between the interconnected load time t.sub.C and the load residence time RT.sub.C can be written as follows:

[00012]$t_C=\frac{A\times {\mathrm{RT}}_C}{c_{\exp }}-t_{\mathrm{CW}}=22.5{\mathrm{RT}}_C-8$

(3) The Solution of the Optimal Number of Operating Columns

[0062] According to the above linear relationship equation, the line t.sub.C -RT.sub.C is drawn in the t-RT coordinate system, as shown in FIG. 6.

[0063] According to the linear equation

[00013]$t=\frac{t_{\mathrm{RR}}-\left(N-3\right)t_{\mathrm{CW}}}{\left(N-2\right)},$

when the number of columns N equals 3, then

[00014]$t=\frac{55-\left(3-3\right)\times 8}{\left(3-2\right)}=55\left(\min \right)$

[0064] In the t-RT coordinate system, line t=55 is drawn.

[0065] As shown in FIG. 6, the RT.sub.C of the intersection of t.sub.C -RT.sub.C and t=55 is 2.8 min. Generally, it is recommended that the single-column residence time in the interconnected load stage of MCC is 2 min-4 min, and this RT.sub.C is within this range. Therefore, the optimal number of operating columns is 3, that is, the three-column continuous chromatography.

(4) The Solution of the Optimal Load Residence Time by Simultaneous Equation Method

[0066] The two linear equations mentioned above are simultaneously solved:

[00015]$\{\begin{array}{c}{t}_C=22.5{\mathrm{RT}}_C-8\\ t_C=55\end{array}$

[0067] The optimal load residence time RT.sub.C,opt is solved and determined to be 2.8 min.

(5) The Solution of the Optimal Load Residence Time by Graphic Method

[0068] According to the equation of the linear relationship between the interconnected load time t.sub.C and the load residence time RT.sub.C mentioned above, the line t.sub.C -RT.sub.C is drawn in the t-RT coordinate system, as shown in FIG. 6.

[0069] According to the linear equation

[00016]$t=\frac{t_{\mathrm{RR}}-\left(N-3\right)t_{\mathrm{CW}}}{\left(N-2\right)},$

when the number of columns N=3, then t=55 min. In the t-RT coordinate system, the line t=55 is drawn.

[0070] The abscissa corresponding to the intersection of the above two lines is the optimal load residence time, which is 2.8 min obtained from FIG. 6.

[0071] Under the conditions of the protein concentration being 4 g/L and the protein breakthrough percentage being 0.7, the three-column MCC experiment is performed with the Praesto® Jetted A50 resin in the load residence time range of 2 min-4 min. The experimental results show that only when the load residence time is 2.8 min, the interconnected load time t.sub.C and the R-R time t.sub.RR are basically equal, and the waiting time is close to 0 min. In this case, the productivity of MCC is 27.6 g/L/h, which is higher than that under other load residence times and is consistent with the optimization results.

Embodiment 3. The Solution for the Maximum Productivity

[0072] The optimal load residence time obtained in Embodiment 2 is substituted into the following formula:

[00017]$P_{C,\mathrm{opt}}=\frac{c_{\exp }}{N\times {\mathrm{RT}}_{C,\mathrm{opt}}}=\frac{4}{3\times 2.8}=0.476g/L/\min =28.6g/L/h$

[0073] The maximum productivity is obtained to be 28.6 g/L/h.

[0074] Under the conditions of the loading protein concentration being 4 g/L, the optimal load residence time being 2.8 min, and the set breakthrough percentage being 0.7, the three-column MCC experiment for protein capture is conducted with the Praesto® Jetted A50 resin. It is found that the loading time t.sub.C and the R-R time t.sub.RR are basically equal. At this time, the productivity of MCC is 27.6 g/L/h, which is close to the predicted maximum productivity of 28.6 g/L/h and higher than the productivity under other conditions. The method of the present invention is confirmed to be effective.

Embodiment 4. The Solution of the Optimal Number of Operating Columns

(1) The Breakthrough Curve Obtained by Experiments

[0075] The Mabselect™ SuRE LX resin from GE Healthcare company is used to pack a 10 ml chromatographic column, the concentration of mAb protein is 8 g/L, and the load residence time is 3 min. The breakthrough experiment is conducted, and the loading is stopped when the breakthrough protein concentration reaches 7.2 g/L. The breakthrough curve is shown in FIG. 7.

(2) The Establishment of the Linear Relationship between the Interconnected Load Time and the Load Residence Time

[0076] When the set breakthrough percentage is 0.75 (that is, 75% breakthrough), the breakthrough curve is integrated to obtain the single-column loading capacity of 85.1 g/L. The R-R time t.sub.RR is 43.75 min and the interconnected wash time t.sub.CW is 6 min after conventional batch chromatography optimization experiment. The above single-column loading capacity, etc., is substituted into the following formula, and the linear relationship between the interconnected load time t.sub.C and the load residence time RT.sub.C can be written as follows:

[00018]$t_C=\frac{A\times {\mathrm{RT}}_C}{c_{\exp }}-t_{\mathrm{CW}}=10.6{\mathrm{RT}}_C-6$

(3) The Solution of the Optimal Number of Operating Columns

[0077] According to the above linear relationship equation, the line t.sub.C -RT.sub.C is drawn in the t-RT coordinate system, as shown in FIG. 8.

[0078] According to the linear equation

[00019]$t=\frac{t_{\mathrm{RR}}-\left(N-3\right)t_{\mathrm{CW}}}{\left(N-2\right)},$

when the number of columns N equals 3, then

[00020]$t=\frac{43.75-\left(3-3\right)\times 6}{\left(3-2\right)}=43.75\left(\min \right)$

[0079] In the t-RT coordinate system, the line t=43.75 is drawn.

[0080] As shown in FIG. 8, the RT.sub.C of the intersection of t.sub.C -RT.sub.C and t=43.75 is 4.7 min. Generally, it is recommended that the single-column residence time in the interconnected load stage of continuous chromatography is 2 min-4 min, and this RT.sub.C is not within this range and is not suitable for the three-column MCC.

[0081] Let the number of columns be N=4, then

[00021]$t=\frac{43.75-\left(4-3\right)\times 6}{\left(4-2\right)}=18.88\left(\min \right)$

[0082] In the t-RT coordinate system, the line t=18.88 is drawn.

[0083] As shown in FIG. 8, the RT.sub.C of the intersection of t.sub.C -RT.sub.C and t=43.75 is 2.3 min, which is within the recommended load residence time range. Therefore, the optimal number of operating columns for this system is 4.

[0084] Under the conditions of the protein concentration being 8 g/L and the protein breakthrough percentage being 0.75, the three-column, four-column, and five-column MCC experiments are performed with the Mabselect SuRE™ LX resin. The maximum productivity of the three modes is 34.7 g/L (the load residence time is 4 min), 52.2 g/L (the load residence time is 2.3 min), and 48 g/L (the load residence time is 2 min), respectively. Specifically, the four-column continuous chromatography has the maximum productivity, which is consistent with the optimization results, so the optimal number of operating columns is verified to be 4.

Embodiment 5. The Solution of the Optimal Load Residence Time

(1) The Breakthrough Curve Obtained by Experiments

[0085] The Mabselect™ SuRE LX resin from GE Healthcare company is used to pack a 10 ml chromatographic column, the concentration of mAb protein is 6 g/L, and the load residence time is 3 min. The breakthrough experiment is conducted, and the loading is stopped when the breakthrough protein concentration reaches 5.4 g/L. The breakthrough curve is shown in FIG. 9.

(2) The Establishment of the Linear Relationship between the Interconnected Load Time and the Load Residence Time

[0086] When the set breakthrough percentage is 0.6 (that is, 60% breakthrough), the breakthrough curve is integrated to obtain the single-column loading capacity of 78.3 g/L. The R-R time t.sub.RR is 43.75 min and the interconnected wash time t.sub.CW is 6 min after optimization by conventional batch chromatography experiment. The above single-column loading capacity, etc., is substituted into the following formula, and the linear relationship between the interconnected load time t.sub.C and the load residence time RT.sub.C can be written as follows:

[00022]$t_C=\frac{A\times {\mathrm{RT}}_C}{c_{\exp }}-t_{\mathrm{CW}}=13.1{\mathrm{RT}}_C-6$

(3) The Solution of the Optimal Number of Operating Columns

[0087] According to the above linear relationship equation, the line t.sub.C -RT.sub.C is drawn in the t-RT coordinate system, as shown in FIG. 10.

[0088] According to the linear equation

[00023]$t=\frac{t_{\mathrm{RR}}-\left(N-3\right)t_{\mathrm{CW}}}{\left(N-2\right)},$

when the number of columns N equals 3, there is:

[00024]$t=\frac{4.375-\left(3-3\right)\times 6}{\left(3-2\right)}=43.75\left(\min \right)$

[0089] In the t-RT coordinate system, the line t=43.75 is drawn.

[0090] As shown in FIG. 10, the RT.sub.C of the intersection of t.sub.C -RT.sub.C and t=43.75 is 3.8 min. Generally, it is recommended that the single-column residence time in the interconnected load stage of continuous chromatography is 2 min-4 min, and this RT.sub.C is within this range. Therefore, the optimal number of operating columns is 3, that is, the three-column continuous chromatography.

(4) The Solution of the Optimal Load Residence Time by Simultaneous Equation Method

[0091] The two linear equations mentioned above are simultaneously solved:

[00025]$\{\begin{array}{c}{t}_C=13.1{\mathrm{RT}}_C-6\\ t_C=43.75\end{array}$

[0092] The optimal load residence time RT.sub.C,opt is solved to be 3.8min.

(5) The Solution of the Optimal Load Residence Time by Graphic Method

[0093] According to the equation of the linear relationship between the interconnected load time t.sub.C and the load residence time RT.sub.C mentioned above, the line t.sub.C -RT.sub.C is drawn in the t-RT coordinate system, as shown in FIG. 10.

[0094] According to the linear equation

[00026]$t=\frac{t_{\mathrm{RR}}-\left(N-3\right)t_{\mathrm{CW}}}{\left(N-2\right)},$

when the number of columns N=3, then t=43.75 min. In the t-RT coordinate system, the line t=43.75 is drawn.

[0095] The abscissa corresponding to the intersection of the above two lines is the optimal load residence time, which is 3.8 min obtained from FIG. 10.

[0096] Under the conditions of the protein concentration being 6 g/L and the protein breakthrough percentage being 0.6, the three-column MCC experiment is performed with the Mabselect™ SuRE LX resin in the load residence time range of 2 min-4 min. The experimental results show that when the load residence time is 3.8 min, the interconnected load time t.sub.C and the R-R time t.sub.RR are basically equal, and the waiting time is close to 0 min. In this case, the productivity of MCC is 30.3 g/L/h, which is higher than that under other load residence times and is consistent with the optimization results.

Embodiment 6. The Solution for the Maximum Productivity

[0097] The optimal load residence time obtained in Embodiment 5 is substituted into the following formula:

[00027]$P_{C,\mathrm{opt}}=\frac{c_{\exp }}{N\times {\mathrm{RT}}_{C,\mathrm{opt}}}=\frac{6}{3\times 3.8}=0.526g/L/\min =31.6g/L/h$

[0098] The maximum productivity is obtained to be 31.6 g/L/h.

[0099] Under the conditions of the loading protein concentration being 6 g/L, the optimal load residence time being 3.8 min, and the set breakthrough percentage being 0.6, the three-column MCC experiment for protein capture is conducted with the Mabselect™ SuRE LX resin. It is found that the loading time t.sub.C and the R-R time t.sub.RR are basically equal. At this time, the productivity of MCC is 30.3 g/L/h, which is close to the predicted maximum productivity of 31.6 g/L/h and higher than the productivity under other conditions. The method of the present invention is confirmed to be effective.

[0100] It should be understood that the exemplary embodiments described herein are illustrative and not restrictive. Although one or more embodiments of the present invention are described in conjunction with the drawings, it should be understood by those of ordinary skill in the art that variations of various forms and details may be made without departing from the spirit and scope of the present invention defined by the claims.