Safe, nontoxic and environment-friendly dewaxing liquid and application thereof

Abstract

The invention discloses a safe, non-toxic and environment-friendly dewaxing liquid for dewaxing a paraffin-embedded tissue slice, and relates to the technical field of biological tissue slice preparation. The specific preparation method comprises the following steps: firstly, weighing Isoparl H, Isoparl M and Isoparl L in a reaction container, uniformly mixing, then weighing amyl alcohol and hexyl alcohol, uniformly mixing, finally adding castor oil, and uniformly mixing, and the components comprise, by weight, 20-30% of Isoparl H, 30-50% of Isoparl M, 10-20% of Isoparl L, 1-10% of castor oil, 1-5% of amyl alcohol and 2-10% of hexyl alcohol. The prepared dewaxing liquid is safe, non-toxic and friendly to human bodies, the health risk caused by long-term exposure when an operator uses a traditional dewaxing agent is reduced, the dewaxing efficiency is high, paraffin can be rapidly and thoroughly removed within 10-20 minutes under the condition of room temperature or slight heating (30-37 DEG C), the tissue form is well kept, high-temperature treatment is not needed, the dewaxing liquid is friendly to the environment, the formula is stable, and the dewaxing liquid is suitable for industrial production. The method is suitable for large-scale production, popularization and application.

Description

Technical Field

[0001] The invention relates to the technical field of biological tissue slice preparation, in particular to a safe, non-toxic and environmentally friendly dewaxing liquid for dewaxing paraffin-embedded tissue slices and its application. Background Art

[0002] Paraffin embedding is the most common method for preserving and sectioning tissue samples in biomedical research, pathological diagnosis, and histological slide preparation. Dewaxing, a crucial step in preparing paraffin-embedded sections, aims to remove paraffin from tissue sections for water-soluble staining or other subsequent processing. Traditional dewaxing methods rely primarily on organic solvents, such as xylene or toluene. While these solvents can effectively dissolve paraffin, they suffer from the following significant drawbacks:

[0003] 1. Significant health hazards: Solvents are highly volatile and have a pungent odor. Long-term exposure to them can cause headaches, nausea, respiratory problems, and even neurological damage.

[0004] 2. Environmental pollution risk: Improper disposal of waste solvents can easily cause soil and water pollution;

[0005] 3. Prominent safety hazards: The solvent has a low flash point and poses a fire hazard.

[0006] Alternative products developed in recent years (such as limonene-based dewaxing agents and petroleum fraction dewaxing agents) have attempted to solve the above problems, but there are still obvious deficiencies in practical applications:

[0007] Inefficient dewaxing: Some alternatives take longer to complete dewaxing;

[0008] Poor user experience: There is a noticeable residual odor;

[0009] Economic and stability deficiencies: Some products are expensive and prone to stratification or deterioration during storage;

[0010] These limitations restrict its widespread application in routine laboratory and clinical settings.

[0011] In summary, there is no dewaxing solution in the prior art that can simultaneously meet the following requirements:

[0012] ①Efficient and thorough removal of paraffin (≤20 minutes);

[0013] ②Safe, non-toxic and no irritating odor;

[0014] ③ Environmentally friendly;

[0015] ④High storage stability;

[0016] ⑤ The cost is controllable and suitable for large-scale application. Summary of the Invention

[0017] Technical problems solved

[0018] This invention addresses the issues of high toxicity, strong pungent odor, flammability, and environmental pollution associated with traditional dewaxing agents (such as xylene). It provides a novel, safe, non-toxic, environmentally friendly, and efficient dewaxing solution for dewaxing paraffin-embedded biological tissue sections. Through the synergistic action of specific components, this dewaxing solution ensures thorough dewaxing of tissue samples while significantly minimizing the impact on operator health and the environment.

[0019] Technical Solution

[0020] The environmentally friendly dewaxing liquid is composed of the following components in percentage by weight:

[0021] IsoparlH: 20-30%

[0022] IsoparlM: 30-50%

[0023] IsoparlL: 10-20%

[0024] Castor oil: 1-10%

[0025] Pentanol: 1-5%

[0026] Hexanol: 2-10%

[0027] The preparation method includes three steps:

[0028] 1. Mix Isoparl H, Isoparl M, and Isoparl L evenly;

[0029] 2. Mix pentanol and hexanol evenly and add them into the above system;

[0030] 3. Finally, add castor oil and mix until homogeneous.

[0031] Component functions and synergistic mechanisms

[0032] Isoparl series solvents (high purity isoparaffins):

[0033] 1) Low toxicity, low odor, suitable volatilization speed and strong paraffin solubility;

[0034] 2) Balancing the paraffin dissolution efficiency and low volatility by compounding isoparaffins with different carbon chain lengths.

[0035] ·castor oil:

[0036] 1) Acts as a natural lubricant to prevent tissue sections from drying out during the dewaxing process;

[0037] 2) Enhance the stability of dewaxing liquid and extend its service life.

[0038] Pentanol and Hexanol:

[0039] 1) As a co-solvent, it improves the penetration and dissolution rate of paraffin;

[0040] 2) Gently protect tissue structure to avoid degeneration or damage.

[0041] Core Advantages

[0042] 1. Safe and non-toxic:

[0043] oAll components are low-irritation and low-toxic materials, which are friendly to the human body and reduce the health risks of operators exposed to long-term exposure.

[0044] 2. Efficient dewaxing:

[0045] Completely remove paraffin wax within 10–20 minutes at room temperature or slightly warmed (30–37°C).

[0046] oNo high temperature treatment is required, and the tissue morphology integrity is well maintained.

[0047] 3. Environmentally friendly:

[0048] oThe solvent is easily degraded after use, meeting the requirements of biomedical research and clinical laboratories for green and sustainable materials.

[0049] 4. Outstanding stability:

[0050] oThe formula is stable and can be stored at room temperature for more than one year without stratification or deterioration, making it suitable for large-scale production and application. BRIEF DESCRIPTION OF THE DRAWINGS

[0051] Figure 1 This is a photograph of a slice made using the environmentally friendly dewaxing solution of the present invention (the slice is not dewaxed).

[0052] Figure 2 This is a photo of a section after dewaxing using the environmentally friendly dewaxing solution of the present invention. DETAILED DESCRIPTION

[0053] To verify the applicability of the environmentally friendly dewaxing solution of the present invention in histological processing, the specific application operations thereof in the wax impregnation process of tissue blocks and HE staining of paraffin sections are introduced below.

[0054] Example 1: Tissue block dehydration, clearing, and wax impregnation process

[0055] After tissue fixation, the dewaxing solution of the present invention is used instead of xylene for transparent treatment. The specific steps are as follows:

[0056] 1) Fix with 10% neutral formalin for 2 hours, changing once and repeating for 2 hours;

[0057] 2) Dehydration in 80% ethanol, 95% ethanol (twice), and anhydrous ethanol (twice), 45 minutes each step;

[0058] 3) using the dewaxing solution of the present invention to make it transparent twice, each time for 5-10 minutes;

[0059] 4) Immerse in 65°C paraffin wax twice (60 minutes each) and a third time (120–180 minutes).

[0060] The experimental results show that the tissue treated with the dewaxing solution of the present invention has good transparency, is fully waxed, and has excellent section quality, making it suitable for subsequent pathological preparation and staining analysis.

[0061] Example 2: Paraffin section HE staining and dewaxing process

[0062] The environmentally friendly dewaxing solution of the present invention is used to dewax and transparentize conventional paraffin sections, and the specific steps are as follows:

[0063] 1) Dewaxing the paraffin sections in the dewaxing solution of the present invention for 5-10 minutes;

[0064] 2) Replace the solution and continue dewaxing for 5–10 minutes to ensure that the paraffin is completely dissolved;

[0065] 3) The sections were rehydrated in a series of ethanol (2–3 minutes), 95% ethanol (1–2 minutes), and 80% ethanol (1–2 minutes).

[0066] 4) Rinse with water for 1–2 minutes;

[0067] 5) Perform routine HE staining (steps omitted);

[0068] 6) After dyeing, dehydration treatment is carried out in sequence (steps omitted);

[0069] 7) performing a first clearing treatment using the dewaxing solution of the present invention for 2-5 minutes;

[0070] 8) Replace the solution and perform a second clearing for 2–5 minutes;

[0071] 9) Finally, perform the sealing operation.

[0072] In the above process, the tissue structure is intact, the staining is clear, the dewaxing is thorough, and there are no residual wax spots. Compared with the traditional use of xylene, the effect is equivalent to or even better, and the risk of odor irritation and toxicity is significantly reduced.

[0073] In summary, the dewaxing solution of the present invention can be widely used in multiple steps of tissue dewaxing, clearing and wax impregnation, and has good application value in replacing traditional toxic solvents.

Claims

1. A safe, non-toxic and environmentally friendly dewaxing liquid, characterized in that: The following components are included by weight percentage: IsoparlH 20-30%, IsoparlM 30~50%, Isopar 10-20%, Castor oil 1-10%, Pentanol 1-5%, Hexanol 2-10%.

2. The environmentally friendly dewaxing liquid according to claim 1, characterized in that: Isoparl H, Isoparl M and Isoparl L are high-purity isoparaffin solvents.

3. The environmentally friendly dewaxing liquid according to claim 1, characterized in that: The castor oil is a natural lubricant and is used to prevent tissue sections from drying out and cracking and to improve the stability of the dewaxing solution.

4. The environmentally friendly dewaxing liquid according to claim 1, characterized in that: Pentanol and hexanol are used as co-solvents to enhance the solvent's ability to penetrate and dissolve paraffin, accelerate the dewaxing process, and gently protect the tissue structure to avoid tissue degeneration or damage.

5. A method for preparing an environmentally friendly dewaxing solution as claimed in any one of claims 1 to 4, characterized in that The following steps are involved: a. Mix IsoparlH, Isoparl M, and Isoparl L; b. The pentanol and hexanol were mixed and then added to the mixture obtained in step a; c. Add castor oil and mix well.

6. Use of the environmentally friendly dewaxing solution according to any one of claims 1 to 4 in the dewaxing, clearing or wax-immersion treatment of paraffin-embedded tissue sections.

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