How to choose & Use a spectrophotometer (Simple Version)

I. Core Selection Factors

1. Define Testing Requirements

  • Wavelength range: Standard 190–1100nm (budget option: 325–1000nm)
  • Sample type: Liquid (standard cuvette) / Solid (integrating sphere) / Micro-volume (micro cell)
  • Accuracy: 1–2nm bandwidth for qualitative analysis; <0.5nm for quantitative

2. Key Configuration Comparison

Feature Basic/Teaching Research-Grade Use Case
Light Source Deuterium + Tungsten Xenon (longer lifespan) Continuous operation
Detector Silicon photodiode PMT (10× sensitivity) Trace analysis
Optical System Grating (1200 lines/mm) Double monochromator Stray light reduction
Data Interface USB/local storage LIMS integration GMP labs

3. Special Functions

  • Kinetics: Time-scan (min. interval: 0.1s)
  • Multi-component: Multi-wavelength/full-spectrum scan
  • Ultra-trace: Fiber optic probe (≤50μL volume)

II. International vs. HINOTEK Comparison

Parameter International Brand Chinese Spectrophotometer (HINOTEK)
Price Range US$10000.00~US$80000.00 <US$300.00 (721G Visible Spectrophotometer)
<US$800.00 (752N UV-Visible Spectrophotometer)
<US$2000.00 (L7 Double Beam UV-Visible Spectrophotometer)
Wavelength Accuracy ±0.1nm ±0.2–0.5nm
Stray Light <0.01%T (220nm) <0.1%T
Scan Speed 5000nm/min 2000nm/min
Recommended Use Pharmacopeia/research Teaching/routine QC

III. Sample Preparation & Measurement Tips

1. Liquid Samples

  • Solvent: Ensure transparency at target wavelength (e.g., quartz cuvette for 260nm)
  • Concentration: Optimal absorbance 0.2–0.8
  • Debubbling: Settle or low-speed centrifugation

2. Cuvette Guidelines (View HINOTEK cell)

Material Wavelength Range Notes
Quartz 190–2500nm Avoid hydrofluoric acid
Glass 340–2500nm Resists strong alkalis
Plastic 380–780nm Single-use only

3. Special Samples

  • Turbid samples: Centrifuge or filter (0.22μm)
  • High absorbance: Dilute or use short-path cuvette (e.g., 1mm)

IV. Instrument Optimization

1. Parameter Setup

  • Scan mode: Fixed wavelength (quantitative) / Full spectrum
  • Bandwidth: 2nm (routine); <1nm (sharp peaks)

2. Baseline Correction

  • Blank solvent calibration (mandatory for solvent changes)
  • Dual-beam compensation (reduces solvent noise)

3. Error Prevention

  • Align cuvette markings (frosted side toward light)
  • Warm up for 30 minutes before use
  • Dilute high-concentration samples (A >1.0 causes nonlinearity)

V. Data Analysis & Validation

1. Quantitative Workflow

  • Calibration curve: 5–7 points, R² >0.999, QC verification
  • Multi-component: Ensure Δλ >30nm to avoid overlap

2. Spectral Peaks

  • Proteins: 280nm (tryptophan/tyrosine)
  • DNA/RNA: 260nm
  • Aromatics: Multi-peaks (200–300nm)

3. Method Validation

  • LOD/LOQ: S/N = 3/10
  • Precision: RSD <1% (6 replicates)

VI. Maintenance & Troubleshooting

1. Routine Care

Frequency Task Standard
Daily Clean cuvette windows No fingerprints/scratches
Weekly Light source energy check >80% initial output
Quarterly Wavelength calibration Holmium oxide standard

2. Common Issues

Symptom Cause Solution
Noisy baseline Lamp aging/power fluctuation Replace lamp/use stabilizer
Drifting readings Cuvette contamination Ultrasonic clean (1% HNO₃)
Wavelength shift Grating misalignment Calibrate with Hg lamp

3. Component Lifespan

  • Deuterium lamp: 1K–2K hours (avoid frequent on/off)
  • Tungsten lamp: 5K hours
  • Cuvettes: Quartz (2 years); glass (1 year)

VII. Safety & Compliance

  • UV protection: Close sample compartment; wear UV-blocking goggles
  • Chemical safety: Use sealed cells for corrosive solvents; segregate waste
  • Electrical: Wait 5 minutes after power loss before restarting

Typical Applications

  1. Protein quantification: Bradford assay (595nm) – avoid detergent interference
  2. Water COD testing: 620nm (dichromate method) or UV 254nm (correlation required)
  3. Nanoparticle analysis: Full scan + Mie theory sizing

Pro Tip: Proper blank correction and cuvette handling eliminate 80% of errors. Establish SOPs, especially for pharmacopeia compliance (wavelength accuracy, absorbance repeatability).

Need more details? Visit our Ultimate UV-Vis Spectrophotometer Buying Guide: From Application Needs to Investment Decisions.

For any inquiries or purchasing needs, please contact the HINOTEK professional team at [email protected].

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