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Cell lysate, method for extracting intracellular protein, method for preparing Toxoplasma antigen, and kit

A technology of cell lysate and Toxoplasma gondii, which is applied in peptide preparation methods, chemical instruments and methods, and biological testing, and can solve problems such as low target protein content, poor thermal stability of antigens, and unfavorable long-term storage

Inactive Publication Date: 2018-07-13
SHENZHEN NEW INDS BIOMEDICAL ENG
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] The lysate used in the prior art still has a bottleneck in the extraction of intracellular proteins, the amount of extracted target protein is low, and many proteins are unstable and easy to degrade, resulting in low detection sensitivity and specificity for in vitro diagnostic items, and diagnostic reagents The antigen used in the box or the antigen after coating has poor thermal stability, which is not conducive to long-term storage

Method used

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  • Cell lysate, method for extracting intracellular protein, method for preparing Toxoplasma antigen, and kit
  • Cell lysate, method for extracting intracellular protein, method for preparing Toxoplasma antigen, and kit
  • Cell lysate, method for extracting intracellular protein, method for preparing Toxoplasma antigen, and kit

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Effect test

Embodiment 1

[0060] 1) Take a frozen strain of Toxoplasma gondii (200 μL), immediately place it at 37°C and thaw it completely within 1 min,

[0061] 2) Inject the thawed Toxoplasma strain into the abdominal cavity of the mouse with a 1mL syringe,

[0062] 3) After 3-4 days, the mice became ill (hair erection, dyspnea, curled up and eyes closed, and abdomen swollen), and they were killed by decapitation.

[0063] 4) Quickly put in 75% alcohol for surface disinfection of the whole body,

[0064] 5) Take out the mouse and fix it, cut off the abdominal skin to expose the peritoneum,

[0065] 6) Use a sterile syringe (5mL) to extract 2-3mL 0.1M PBS (pH 7.2), inject it into the abdominal cavity (without damaging internal organs), then gently massage the abdomen to mix the PBS and peritoneal fluid evenly,

[0066] 7) Collect the peritoneal fluid, put it in a sterile test tube, and repeatedly take PBS for intraperitoneal injection until the withdrawn peritoneal fluid becomes clear,

[0067] 8)...

Embodiment 2

[0084] Embodiment 2-replacing Tween-20 in embodiment 1 with Triton X-100, all the other are consistent with embodiment 1.

Embodiment 3

[0085] Embodiment 3-replacing the sucrose in embodiment 1 with maltose, all the other are consistent with embodiment 1.

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Abstract

The invention relates to extraction, preparation and detection of biological substances, in particular to a cell lysate, a method for extracting intracellular proteins such as a Toxoplasma membrane antigen by using the cell lysate, and a kit employing the Toxoplasma membrane antigen, belonging to the field of in-vitro diagnostic reagents. The cell lysate of the invention includes amino acids, denaturants, carbohydrates and alcohols. Compared with the prior art, the invention has the advantages that intracellular proteins are fully released and a recovery rate is increased since a novel formulafor the lysate is adopted, so the sensitivity and specificity of clinical diagnosis and detection are improved. At the same time, the novel lysate of the nvention promotes tight folding of proteins,stabilizes the conformations of proteins, enhances the thermal stability of proteins, and ensures stable storage of detection reagents.

Description

technical field [0001] The invention belongs to the field of in vitro diagnostic reagents, relates to the extraction, preparation and detection of biological substances, in particular to a cell lysate, a method for extracting intracellular proteins, such as toxoplasma membrane antigen, using the cell lysate, and using the toxoplasma A kit for entomoantigens. Background technique [0002] The extraction of intracellular proteins has a wide range of applications in the field of in vitro diagnostic antigen preparation, especially some key antigen raw material proteins, which exist in a small amount in cells, and most of them are low-abundance protein antigens, so the extraction of intracellular proteins is particularly important. Importantly, extraction of high concentrations of immunologically functional proteins is a critical part of the in vitro diagnostic process. [0003] The extraction process of intracellular protein generally includes the preparation of a lysate with a...

Claims

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Application Information

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IPC IPC(8): C07K1/14G01N33/531G01N33/68G01N33/569
CPCC07K1/145G01N33/531G01N33/56905G01N33/6854G01N2469/20
Inventor 饶微刘振华何海华
Owner SHENZHEN NEW INDS BIOMEDICAL ENG
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