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Virus and antigen purification and conjugation

A virus and antigen technology, applied in the field of virus and antigen purification and coupling, can solve the problem of insufficient removal of impurities

Active Publication Date: 2021-03-26
켄터키바이오프로세싱인코포레이티드
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, current methods of purifying virus will not adequately remove all or even sufficient amounts of impurities, including but not limited to those present in plant extracts, and will not yet adequately produce purified virus

Method used

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  • Virus and antigen purification and conjugation
  • Virus and antigen purification and conjugation
  • Virus and antigen purification and conjugation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0095] Embodiment 1—purification of icosahedral red clover mosaic virus

[0096] As a known technique for detecting various proteins in a mixture, image 3 The Western Blot provided in , which shows figure 2 The successful purification of the icosahedral red clover mosaic virus is shown. Similarly, Figure 5 Western blot in Figure 4 The successful purification of the icosahedral red clover mosaic virus is shown. Both viruses were purified according to the embodiments described herein. The target protein is extracted from the tissue according to known detection techniques. The proteins of the sample are then separated using gel electrophoresis according to their isoelectric point, molecular weight, charge, or various combinations of these factors. The samples are then loaded into individual lanes in the gel, with one lane reserved for a "ladder" containing a mixture of known proteins of defined molecular weight. For example in image 3 In , lane 12 serves as a ladde...

Embodiment 2

[0099] Example 2—Purification of Rod-Shaped TMV

[0100] Image 6 shows purified rod-shaped TMV, Figure 7 A viral purification platform for TMV to achieve this purification is shown within the scope of the various embodiments and alternatives disclosed herein. and image 3 with Figure 5 similar, Figure 7 Shows the purity of the virus product after each step of the virus purification platform is completed. After the final purification step, the resulting product is a highly purified virus product, with Figure 7 The clearly visible bands in lane 13 are consistent.

[0101] Thus, the virus purification platform of the present invention has successfully purified every virus for which the inventors have applied these methods, including icosahedral and baculoviruses, and the platform is expected to reproducibly and consistently purify almost Viruses of any type, if not all types.

[0102] Production and purification of recombinant antigens

[0103] Table 2 and Fig...

Embodiment 3、4、5 and 6

[0116] Examples 3, 4, 5 and 6—H5 rHA, H7 rhA, WNV rDIII and LFV rGP1 / 2

[0117] Such as Figure 12 As shown, antigen purification platforms according to various embodiments and alternatives have successfully purified H5rHA, H7 rhA, WNV rDIII and LFV rGP1 / 2. Figure 12 Contains two images taken from the conclusion of the antigen purification platform: the left image contains an SDS Page gel indicating the purity of the viral vector TMV NtK (where NtK is an abbreviation for N-terminal lysine) and influenza antigens, and the right image contains a Western blot , indicating immunoreactivity to West Nile and Lassa fever antigens. Such as Figure 12 Each antigen product is highly pure, as shown by the clearly visible bands in . Thus, the antigen purification platform according to various embodiments and alternatives consistently purifies each type of antigen on a commercial scale in a manner compliant with cGMP regulations. In the same way, the platform promises to rep...

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Abstract

Disclosed herein are methods and exemplary compositions associated with virus purification, antigen purification, and conjugation of virus and proteins (e.g., antigen) to form vaccines for delivery ofimmunological and other therapeutic agents, exemplary aspects of which may include harvesting viral and antigenic substances from source organisms; a purification platform comprising chemical separation and size-difference separation for the removal of contaminants, debris and impurities from the viral and protein (e.g. antigenic, including influenza hemagglutinin antigens) substances, as well astheir concentration and collection; and a conjugation platform providing activation of the virus at a pH that increases binding rate and binding propensity between the virus and the protein, whereinembodiments related to the conjugation platform include controlling the ratio of virus to protein.

Description

[0001] U.S. Nonprovisional Patent Application [0002] Cross References to Related Applications [0003] This international patent application claims the benefit and priority of U.S. Provisional Patent Application Serial No. 62 / 683,865, filed June 12, 2018, the contents of which are incorporated herein by reference in their entirety. technical field [0004] Embodiments described herein include the use of a multi-set process for the production of highly purified recombinant viruses as antigen carriers, and further various embodiments relate to vaccine production using purified viruses and purified antigens. Background technique [0005] Viruses have nucleic acid molecules in a protein coat and only replicate inside the living cells of other organisms. Generally considered harmful, a wide variety of viruses are capable of infecting all types of life forms such as humans, livestock and plants. But on the positive side, there is growing interest in using viruses for a variety...

Claims

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Application Information

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IPC IPC(8): A01K67/027A61P37/04C07K14/00C07K14/11C12N5/0793C12N5/0797
CPCB01D15/34B01D15/361B01D15/3809B01D15/3847A61K47/6901C12N2760/16122C12N2760/16222C12N2770/00051C12N2770/00063C12N2770/40051C12N2770/40063C12N2770/00043C12N2770/00034A61K2039/5258A61K2039/60A61K2039/5252C12N2770/24134C12N2760/10034A61K39/12C12N7/00C07K1/16C07K1/18A61K39/145C12N2770/40061C07K1/22A61K38/36A61K39/0001C12N2770/00061A61K2039/5254C12N15/86C07K14/005C07K1/165C12N2760/16134C12N2760/16234
Inventor L·博尔登S·D·修梅J·莫顿G·波格B·布拉彻H·A·海登C·A·辛普森N·帕坦J·W·舍普赫德K·斯沃普
Owner 켄터키바이오프로세싱인코포레이티드
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