📚 A-Level OCR Biology: Practical Skills Guide | A-Level OCR 生物：实验操作指南
Mastering practical techniques is essential for success in OCR A-Level Biology. This guide covers the key experimental procedures, from microscopy and biochemical tests to enzyme kinetics and aseptic technique, helping you build confidence for practical assessments and written exams.
掌握实验操作技术是OCR A-Level生物考试成功的关键。本指南涵盖从显微镜使用、生化测试到酶动力学和无菌操作的核心实验步骤，助你提升实践评估和笔试的信心。
1. Using a Microscope and Calibrating the Eyepiece Graticule | 使用显微镜与校准目镜测微尺
To measure cell dimensions accurately, you must calibrate the eyepiece graticule using a stage micrometer. Place the stage micrometer on the stage, focus with the lowest power objective, and align the scales. Count how many graticule divisions correspond to a known length on the micrometer, e.g. 10 graticule divisions = 0.1 mm. Calculate the length of one graticule division in micrometres for that objective, and repeat for each objective lens to produce a calibration table.
要精确测量细胞大小，必须使用镜台测微尺校准目镜测微尺。将镜台测微尺置于载物台，以低倍物镜对焦后对齐标尺。数出测微尺上已知长度（如10小格=0.1 mm）所对应的目镜测微尺格数，计算该物镜下每格长度（µm），并对所有物镜重复此操作以建立校准表。
The calibration factor is used when measuring a specimen: replace the stage micrometer with the specimen slide, then use the eyepiece graticule to count the number of divisions across the cell. Multiply the division count by the calibration factor to obtain the actual cell size. Always record the total magnification and which objective lens was used for the measurement.
校准后的换算因子可用于样本测量：将镜台测微尺换成标本玻片，利用目镜测微尺数出跨越细胞所占的格数，再乘以校准因子即得实际尺寸。务必记录总放大倍数及所用物镜。
Actual size = Number of graticule divisions × Calibration factor (µm per division)
实际大小 = 目镜测微尺格数 × 校准因子（每格微米数）
2. Preparing Temporary Mounts and Staining | 制作临时装片与染色
A wet mount is made by placing a drop of water or appropriate stain onto a clean microscope slide. Use forceps to transfer the specimen, such as onion epidermis or cheek cells, into the drop. Hold a coverslip at a 45° angle and lower it gently to avoid trapping air bubbles. Excess liquid can be removed with filter paper.
制作水封片时，在洁净载玻片上滴加一滴水或适宜染液，用镊子将洋葱表皮或口腔上皮细胞等标本放入液滴中。以45°角持盖玻片轻轻放下，以免产生气泡，多余液体用滤纸吸干。
Staining greatly improves contrast. Iodine–KI solution stains starch blue-black and nuclei brown, while methylene blue stains nuclei dark blue. Stains can be added before lowering the coverslip, or by the irrigation method: place a drop of stain at one edge of the coverslip and draw it through by touching filter paper to the opposite edge.
染色可大幅提升反差。碘-碘化钾染液使淀粉呈蓝黑色、细胞核呈褐色；亚甲蓝则使细胞核呈深蓝色。可在加盖玻片前滴加染液，也可采用引流法：在盖玻片一侧滴加染液，另一侧用滤纸引流。
3. Biochemical Tests for Biological Molecules | 生物分子的生化测试
The Benedict’s test identifies reducing sugars. Add excess Benedict’s reagent to the sample and heat in a water bath at ≥80 °C. A colour change from blue through green, yellow, orange to brick-red precipitate confirms the presence and relative amount of reducing sugar. For non-reducing sugars such as sucrose, first boil the sample with dilute HCl to hydrolyse, neutralise with NaHCO₃, then perform the Benedict’s test; an orange-red precipitate indicates non-reducing sugar originally present.
本尼迪克特测试可鉴定还原糖。加入过量本尼迪克特试剂后在≥80 °C水浴加热，溶液由蓝变绿、黄、橙至砖红色沉淀，证实还原糖存在并可粗略定量。检测蔗糖等非还原糖时，先用稀盐酸煮沸水解，再用碳酸氢钠中和，然后进行本尼迪克特测试，出现橙红色沉淀则说明原样中含有非还原糖。
The iodine test for starch: add a few drops of iodine–KI solution; a blue-black colour is positive. The emulsion test for lipids: shake the sample with ethanol, pour the liquid into water; a milky-white emulsion indicates lipids. The Biuret test for proteins: add Biuret reagent (NaOH followed by CuSO₄) and look for a violet colour change; a negative control remains blue.
淀粉的碘测试：加入碘-碘化钾溶液，显蓝黑色为阳性。脂质的乳浊测试：将样品与乙醇振摇后倒入水中，乳白色乳浊液证实脂质。蛋白质的双缩脲测试：加入双缩脲试剂（氢氧化钠与硫酸铜），溶液变紫色为阳性，阴性对照保持蓝色。
4. Investigating Enzyme Activity: Temperature and pH | 酶活性探究：温度与pH
To investigate the effect of temperature on trypsin, set up water baths across a range (e.g. 10–60 °C). Mix trypsin solution with milk suspension in a cuvette, place it in the water bath, and use a colorimeter to record the time taken for the absorbance to fall by a standard amount, or until the mixture clears. All solutions must be equilibrated at the target temperature before mixing.
探究温度对胰蛋白酶的影响时，设置10–60 °C的系列水浴。将胰蛋白酶溶液与牛奶悬浊液在比色皿中混合，置于水浴，用比色计记录吸光度下降固定值所需时间或直至混合物澄清的时间。所有溶液须在混合前于目标温度下平衡。
Rate of reaction is expressed as 1/time. Plotting rate against temperature yields a curve that rises to an optimum, then falls sharply owing to denaturation of the enzyme’s tertiary structure. For pH, use buffer solutions (pH 2, 4, 6, 7, 8, 10) and maintain a constant optimum temperature. The rate vs pH graph is bell-shaped, peaking at the enzyme’s optimum pH.
反应速率以1/时间表示，绘制速率-温度曲线会呈现升至最适温度后因酶的三级结构变性而急剧下降的趋势。探究pH时使用缓冲液（pH 2、4、6、7、8、10）并保持恒温，速率-pH曲线呈钟形，峰值出现在酶的最适pH处。
5. Measuring the Rate of Photosynthesis using Algal Balls | 使用藻球测量光合作用速率
Immobilise unicellular algae such as Chlorella in sodium alginate beads. Use hydrogencarbonate indicator, which is red at atmospheric CO₂ level, turns yellow when CO₂ concentration increases, and purple when CO₂ decreases. Place a standardised number of algal balls in vials containing indicator and expose to various light intensities, e.g. by varying the distance from a lamp. Include a control vial without algae.
将小球藻等单细胞藻类用海藻酸钠固定成珠。碳酸氢盐指示液在大气CO₂浓度时呈红色，CO₂升高变黄，CO₂降低变紫。将定量藻球放入盛有指示液的小瓶中，通过改变灯距获得不同光强，并设置无藻球的对照瓶。
Photosynthesis removes CO₂, turning the indicator purple; the rate of colour change reflects the photosynthetic rate. Use a colorimeter to measure absorbance at timed intervals, or record the time taken for the indicator to shift from red to purple. Plot absorbance or 1/time against light intensity to produce a response curve, highlighting the light compensation point and saturation.
光合作用消耗CO₂使指示液变为紫色，颜色变化速率对应光合速率。用比色计定时测定吸光度，或记录指示液从红变紫所需时间。绘制吸光度或1/时间对光强的曲线，可体现光补偿点与光饱和现象。
6. Investigating Respiration in Yeast | 探究酵母的呼吸作用
For anaerobic respiration, set up a flask containing yeast suspension and glucose solution, connected to a gas syringe. Place the apparatus in a water bath at 30 °C. As yeast ferments, it produces CO₂; record the volume of gas collected at 1-minute intervals for 5–10 minutes. Rate of fermentation can be calculated as volume of CO₂ per minute.
探究无氧呼吸时，将酵母悬浮液与葡萄糖溶液置于烧瓶并连接气体注射器，于30 °C水浴中保温。酵母发酵产生CO₂，每隔1分钟记录收集的气体体积，持续5–10分钟。发酵速率以每分钟产CO₂体积计。
Aerobic respiration can be measured with a respirometer. Place germinating seeds or yeast in a tube with KOH solution to absorb CO₂. As O₂ is consumed, the manometer fluid moves; the rate of movement indicates oxygen consumption. A thermobarometer tube, with no living material but with KOH, should be used to correct for temperature and pressure fluctuations.
有氧呼吸可用呼吸计测量。将萌发种子或酵母置于含KOH溶液（吸收CO₂）的试管中。消耗O₂时压力计液面移动，移动速率反映耗氧速率。需设置温压计（含KOH但不含生物材料的对照管）以校正温度与气压波动。
7. Separation of Photosynthetic Pigments by Chromatography | 色谱法分离光合色素
Extract pigments by grinding a leaf (e.g. spinach) with a pinch of sand and propanone (acetone) in a mortar. Concentrate the extract and apply a small spot onto a pencil line on TLC plate or chromatography paper. Place the plate in a sealed tank with a solvent mixture such as petroleum ether and propanone (9:1). Allow the solvent to ascend; pigments separate according to their solubility in the solvent and their adsorption to the stationary phase.
将叶片（如菠菜）与少量石英砂和丙酮共同研磨提取色素。浓缩后，在TLC板或色谱纸的铅笔线上点样。将板放入含石油醚与丙酮（9:1）混合液的密闭展缸中，溶剂前沿上升时，色素因溶解度与吸附力差异而分离。
Identify bands by colour and Rf value. Typical Rf values: carotene (orange, ~0.95), pheophytin (grey, ~0.7), chlorophyll a (blue-green, ~0.6), chlorophyll b (yellow-green, ~0.5), xanthophylls (yellow, ~0.3–0.4). Calculate Rf using the formula below. Mark the solvent front immediately after removal.
按颜色与Rf值鉴定色素条带。典型Rf值：胡萝卜素（橙黄，~0.95）、脱镁叶绿素（灰，~0.7）、叶绿素a（蓝绿，~0.6）、叶绿素b（黄绿，~0.5）、叶黄素（黄，~0.3–0.4）。取出后立刻标记溶剂前沿并用下式计算Rf。
Rf = distance moved by spot / distance moved by solvent front
Rf = 色素斑点移动距离 / 溶剂前沿移动距离
8. Determining Water Potential by Measuring Mass Change | 通过质量变化测定水势
Prepare a concentration series of sucrose solutions, e.g. 0.0, 0.2, 0.4, 0.6, 0.8, 1.0 mol dm⁻³. Using a cork borer, cut equal-sized potato cylinders, blot them dry, and record the initial mass. Immerse the cylinders in the solutions for a standardised time (e.g. 30 min). Afterwards, blot and reweigh. Calculate percentage change in mass: [(final mass – initial mass) / initial mass] × 100%.
配制系列蔗糖溶液，如0.0、0.2、0.4、0.6、0.8、1.0 mol dm⁻³。用打孔器切取大小一致的土豆条，吸干表面水分后称初重，浸入各溶液一定时间（如30分钟）。取出吸干后再次称重，计算质量变化百分比。
Plot percentage mass change against sucrose concentration. The point where the line crosses the x-axis (zero mass change) corresponds to the water potential of the potato tissue. Use a conversion table to find the solute potential (ψₛ) of that sucrose concentration, expressed in kPa. The water potential of the tissue equals this ψₛ, assuming the tissue has negligible pressure potential when flaccid.
绘制质量变化百分比对蔗糖浓度的曲线，与x轴交点（质量变化为0）即土豆组织的水势。由换算表查出该浓度蔗糖的溶质势（ψₛ，以kPa表示），组织水势即等于该ψₛ（假设组织质壁分离时压力势为零）。
Ψ = Ψₛ + Ψₚ; for flaccid cells, Ψₚ = 0, so Ψ = Ψₛ
水势 Ψ =
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